Juxtamembrane Shedding of Plasmodium falciparum AMA1 Is Sequence Independent and Essential, and Helps Evade Invasion-Inhibitory Antibodies
Events
Loading … Spinner

Mendeley | Further Information

{"title"=>"Juxtamembrane shedding of plasmodium falciparum AMA1 is sequence independent and essential, and helps evade invasion-inhibitory antibodies", "type"=>"journal", "authors"=>[{"first_name"=>"Anna", "last_name"=>"Olivieri", "scopus_author_id"=>"36882582900"}, {"first_name"=>"Christine R.", "last_name"=>"Collins", "scopus_author_id"=>"7401740087"}, {"first_name"=>"Fiona", "last_name"=>"Hackett", "scopus_author_id"=>"6701535246"}, {"first_name"=>"Chrislaine", "last_name"=>"Withers-Martinez", "scopus_author_id"=>"6603275171"}, {"first_name"=>"Joshua", "last_name"=>"Marshall", "scopus_author_id"=>"9333361900"}, {"first_name"=>"Helen R.", "last_name"=>"Flynn", "scopus_author_id"=>"24480959800"}, {"first_name"=>"J. Mark", "last_name"=>"Skehel", "scopus_author_id"=>"7005850091"}, {"first_name"=>"Michael J.", "last_name"=>"Blackman", "scopus_author_id"=>"7102787691"}], "year"=>2011, "source"=>"PLoS Pathogens", "identifiers"=>{"sgr"=>"84855282762", "pui"=>"364024599", "doi"=>"10.1371/journal.ppat.1002448", "pmid"=>"22194692", "scopus"=>"2-s2.0-84855282762", "issn"=>"15537366", "isbn"=>"1553-7374 (Electronic)\\r1553-7366 (Linking)"}, "id"=>"c1fd7c2e-59c9-3f2c-96b2-29e1f1f84ce0", "abstract"=>"The malarial life cycle involves repeated rounds of intraerythrocytic replication interspersed by host cell rupture which releases merozoites that rapidly invade fresh erythrocytes. Apical membrane antigen-1 (AMA1) is a merozoite protein that plays a critical role in invasion. Antibodies against AMA1 prevent invasion and can protect against malaria in vivo, so AMA1 is of interest as a malaria vaccine candidate. AMA1 is efficiently shed from the invading parasite surface, predominantly through juxtamembrane cleavage by a membrane-bound protease called SUB2, but also by limited intramembrane cleavage. We have investigated the structural requirements for shedding of Plasmodium falciparum AMA1 (PfAMA1), and the consequences of its inhibition. Mutagenesis of the intramembrane cleavage site by targeted homologous recombination abolished intramembrane cleavage with no effect on parasite viability in vitro. Examination of PfSUB2-mediated shedding of episomally-expressed PfAMA1 revealed that the position of cleavage is determined primarily by its distance from the parasite membrane. Certain mutations at the PfSUB2 cleavage site block shedding, and parasites expressing these non-cleavable forms of PfAMA1 on a background of expression of the wild type gene invade and replicate normally in vitro. The non-cleavable PfAMA1 is also functional in invasion. However - in contrast to the intramembrane cleavage site - mutations that block PfSUB2-mediated shedding could not be stably introduced into the genomic pfama1 locus, indicating that some shedding of PfAMA1 by PfSUB2 is essential. Remarkably, parasites expressing shedding-resistant forms of PfAMA1 exhibit enhanced sensitivity to antibody-mediated inhibition of invasion. Drugs that inhibit PfSUB2 activity should block parasite replication and may also enhance the efficacy of vaccines based on AMA1 and other merozoite surface proteins.", "link"=>"http://www.mendeley.com/research/juxtamembrane-shedding-plasmodium-falciparum-ama1-sequence-independent-essential-helps-evade-invasio", "reader_count"=>67, "reader_count_by_academic_status"=>{"Unspecified"=>1, "Professor > Associate Professor"=>5, "Researcher"=>16, "Student > Doctoral Student"=>3, "Student > Ph. D. Student"=>19, "Student > Postgraduate"=>3, "Student > Master"=>11, "Other"=>3, "Student > Bachelor"=>5, "Professor"=>1}, "reader_count_by_user_role"=>{"Unspecified"=>1, "Professor > Associate Professor"=>5, "Researcher"=>16, "Student > Doctoral Student"=>3, "Student > Ph. D. Student"=>19, "Student > Postgraduate"=>3, "Student > Master"=>11, "Other"=>3, "Student > Bachelor"=>5, "Professor"=>1}, "reader_count_by_subject_area"=>{"Unspecified"=>1, "Biochemistry, Genetics and Molecular Biology"=>4, "Agricultural and Biological Sciences"=>48, "Medicine and Dentistry"=>9, "Business, Management and Accounting"=>1, "Pharmacology, Toxicology and Pharmaceutical Science"=>1, "Chemistry"=>2, "Social Sciences"=>1}, "reader_count_by_subdiscipline"=>{"Medicine and Dentistry"=>{"Medicine and Dentistry"=>9}, "Chemistry"=>{"Chemistry"=>2}, "Social Sciences"=>{"Social Sciences"=>1}, "Agricultural and Biological Sciences"=>{"Agricultural and Biological Sciences"=>48}, "Business, Management and Accounting"=>{"Business, Management and Accounting"=>1}, "Biochemistry, Genetics and Molecular Biology"=>{"Biochemistry, Genetics and Molecular Biology"=>4}, "Unspecified"=>{"Unspecified"=>1}, "Pharmacology, Toxicology and Pharmaceutical Science"=>{"Pharmacology, Toxicology and Pharmaceutical Science"=>1}}, "reader_count_by_country"=>{"United States"=>3, "United Kingdom"=>1, "Kenya"=>1, "Germany"=>1, "India"=>1}, "group_count"=>2}

Scopus | Further Information

{"@_fa"=>"true", "link"=>[{"@_fa"=>"true", "@ref"=>"self", "@href"=>"https://api.elsevier.com/content/abstract/scopus_id/84855282762"}, {"@_fa"=>"true", "@ref"=>"author-affiliation", "@href"=>"https://api.elsevier.com/content/abstract/scopus_id/84855282762?field=author,affiliation"}, {"@_fa"=>"true", "@ref"=>"scopus", "@href"=>"https://www.scopus.com/inward/record.uri?partnerID=HzOxMe3b&scp=84855282762&origin=inward"}, {"@_fa"=>"true", "@ref"=>"scopus-citedby", "@href"=>"https://www.scopus.com/inward/citedby.uri?partnerID=HzOxMe3b&scp=84855282762&origin=inward"}], "prism:url"=>"https://api.elsevier.com/content/abstract/scopus_id/84855282762", "dc:identifier"=>"SCOPUS_ID:84855282762", "eid"=>"2-s2.0-84855282762", "dc:title"=>"Juxtamembrane shedding of plasmodium falciparum AMA1 is sequence independent and essential, and helps evade invasion-inhibitory antibodies", "dc:creator"=>"Olivieri A.", "prism:publicationName"=>"PLoS Pathogens", "prism:issn"=>"15537366", "prism:eIssn"=>"15537374", "prism:volume"=>"7", "prism:issueIdentifier"=>"12", "prism:pageRange"=>nil, "prism:coverDate"=>"2011-12-01", "prism:coverDisplayDate"=>"December 2011", "prism:doi"=>"10.1371/journal.ppat.1002448", "citedby-count"=>"26", "affiliation"=>[{"@_fa"=>"true", "affilname"=>"MRC National Institute for Medical Research", "affiliation-city"=>"London", "affiliation-country"=>"United Kingdom"}], "pubmed-id"=>"22194692", "prism:aggregationType"=>"Journal", "subtype"=>"ar", "subtypeDescription"=>"Article", "article-number"=>"e1002448", "source-id"=>"4000151809", "openaccess"=>"1", "openaccessFlag"=>true}

Facebook

  • {"url"=>"http%3A%2F%2Fjournals.plos.org%2Fplospathogens%2Farticle%3Fid%3D10.1371%252Fjournal.ppat.1002448", "share_count"=>0, "like_count"=>0, "comment_count"=>0, "click_count"=>0, "total_count"=>0}

Counter

  • {"month"=>"12", "year"=>"2011", "pdf_views"=>"112", "xml_views"=>"1", "html_views"=>"367"}
  • {"month"=>"1", "year"=>"2012", "pdf_views"=>"141", "xml_views"=>"68", "html_views"=>"191"}
  • {"month"=>"2", "year"=>"2012", "pdf_views"=>"27", "xml_views"=>"0", "html_views"=>"74"}
  • {"month"=>"3", "year"=>"2012", "pdf_views"=>"27", "xml_views"=>"0", "html_views"=>"112"}
  • {"month"=>"4", "year"=>"2012", "pdf_views"=>"17", "xml_views"=>"1", "html_views"=>"57"}
  • {"month"=>"5", "year"=>"2012", "pdf_views"=>"17", "xml_views"=>"0", "html_views"=>"45"}
  • {"month"=>"6", "year"=>"2012", "pdf_views"=>"9", "xml_views"=>"0", "html_views"=>"40"}
  • {"month"=>"7", "year"=>"2012", "pdf_views"=>"8", "xml_views"=>"1", "html_views"=>"40"}
  • {"month"=>"8", "year"=>"2012", "pdf_views"=>"13", "xml_views"=>"5", "html_views"=>"31"}
  • {"month"=>"9", "year"=>"2012", "pdf_views"=>"14", "xml_views"=>"0", "html_views"=>"32"}
  • {"month"=>"10", "year"=>"2012", "pdf_views"=>"8", "xml_views"=>"0", "html_views"=>"37"}
  • {"month"=>"11", "year"=>"2012", "pdf_views"=>"9", "xml_views"=>"0", "html_views"=>"38"}
  • {"month"=>"12", "year"=>"2012", "pdf_views"=>"14", "xml_views"=>"0", "html_views"=>"32"}
  • {"month"=>"1", "year"=>"2013", "pdf_views"=>"9", "xml_views"=>"0", "html_views"=>"69"}
  • {"month"=>"2", "year"=>"2013", "pdf_views"=>"10", "xml_views"=>"2", "html_views"=>"189"}
  • {"month"=>"3", "year"=>"2013", "pdf_views"=>"7", "xml_views"=>"0", "html_views"=>"25"}
  • {"month"=>"4", "year"=>"2013", "pdf_views"=>"7", "xml_views"=>"0", "html_views"=>"119"}
  • {"month"=>"5", "year"=>"2013", "pdf_views"=>"13", "xml_views"=>"0", "html_views"=>"71"}
  • {"month"=>"6", "year"=>"2013", "pdf_views"=>"3", "xml_views"=>"0", "html_views"=>"22"}
  • {"month"=>"7", "year"=>"2013", "pdf_views"=>"7", "xml_views"=>"1", "html_views"=>"25"}
  • {"month"=>"8", "year"=>"2013", "pdf_views"=>"7", "xml_views"=>"0", "html_views"=>"30"}
  • {"month"=>"9", "year"=>"2013", "pdf_views"=>"8", "xml_views"=>"0", "html_views"=>"33"}
  • {"month"=>"10", "year"=>"2013", "pdf_views"=>"1", "xml_views"=>"1", "html_views"=>"63"}
  • {"month"=>"11", "year"=>"2013", "pdf_views"=>"8", "xml_views"=>"1", "html_views"=>"87"}
  • {"month"=>"12", "year"=>"2013", "pdf_views"=>"9", "xml_views"=>"1", "html_views"=>"74"}
  • {"month"=>"1", "year"=>"2014", "pdf_views"=>"14", "xml_views"=>"0", "html_views"=>"88"}
  • {"month"=>"2", "year"=>"2014", "pdf_views"=>"6", "xml_views"=>"0", "html_views"=>"60"}
  • {"month"=>"3", "year"=>"2014", "pdf_views"=>"3", "xml_views"=>"3", "html_views"=>"153"}
  • {"month"=>"4", "year"=>"2014", "pdf_views"=>"6", "xml_views"=>"1", "html_views"=>"140"}
  • {"month"=>"5", "year"=>"2014", "pdf_views"=>"4", "xml_views"=>"0", "html_views"=>"58"}
  • {"month"=>"6", "year"=>"2014", "pdf_views"=>"5", "xml_views"=>"0", "html_views"=>"53"}
  • {"month"=>"7", "year"=>"2014", "pdf_views"=>"3", "xml_views"=>"0", "html_views"=>"43"}
  • {"month"=>"8", "year"=>"2014", "pdf_views"=>"2", "xml_views"=>"2", "html_views"=>"67"}
  • {"month"=>"9", "year"=>"2014", "pdf_views"=>"3", "xml_views"=>"0", "html_views"=>"54"}
  • {"month"=>"10", "year"=>"2014", "pdf_views"=>"4", "xml_views"=>"1", "html_views"=>"38"}
  • {"month"=>"11", "year"=>"2014", "pdf_views"=>"4", "xml_views"=>"1", "html_views"=>"84"}
  • {"month"=>"12", "year"=>"2014", "pdf_views"=>"8", "xml_views"=>"2", "html_views"=>"38"}
  • {"month"=>"1", "year"=>"2015", "pdf_views"=>"1", "xml_views"=>"0", "html_views"=>"27"}
  • {"month"=>"2", "year"=>"2015", "pdf_views"=>"6", "xml_views"=>"0", "html_views"=>"20"}
  • {"month"=>"3", "year"=>"2015", "pdf_views"=>"6", "xml_views"=>"0", "html_views"=>"26"}
  • {"month"=>"4", "year"=>"2015", "pdf_views"=>"6", "xml_views"=>"1", "html_views"=>"35"}
  • {"month"=>"5", "year"=>"2015", "pdf_views"=>"5", "xml_views"=>"0", "html_views"=>"30"}
  • {"month"=>"6", "year"=>"2015", "pdf_views"=>"2", "xml_views"=>"0", "html_views"=>"49"}
  • {"month"=>"7", "year"=>"2015", "pdf_views"=>"4", "xml_views"=>"0", "html_views"=>"21"}
  • {"month"=>"8", "year"=>"2015", "pdf_views"=>"5", "xml_views"=>"0", "html_views"=>"20"}
  • {"month"=>"9", "year"=>"2015", "pdf_views"=>"5", "xml_views"=>"0", "html_views"=>"25"}
  • {"month"=>"10", "year"=>"2015", "pdf_views"=>"4", "xml_views"=>"0", "html_views"=>"26"}
  • {"month"=>"11", "year"=>"2015", "pdf_views"=>"3", "xml_views"=>"0", "html_views"=>"15"}
  • {"month"=>"12", "year"=>"2015", "pdf_views"=>"2", "xml_views"=>"0", "html_views"=>"15"}
  • {"month"=>"1", "year"=>"2016", "pdf_views"=>"9", "xml_views"=>"0", "html_views"=>"13"}
  • {"month"=>"2", "year"=>"2016", "pdf_views"=>"14", "xml_views"=>"0", "html_views"=>"12"}
  • {"month"=>"3", "year"=>"2016", "pdf_views"=>"10", "xml_views"=>"0", "html_views"=>"35"}
  • {"month"=>"4", "year"=>"2016", "pdf_views"=>"4", "xml_views"=>"0", "html_views"=>"39"}
  • {"month"=>"5", "year"=>"2016", "pdf_views"=>"1", "xml_views"=>"0", "html_views"=>"11"}
  • {"month"=>"6", "year"=>"2016", "pdf_views"=>"2", "xml_views"=>"0", "html_views"=>"9"}
  • {"month"=>"7", "year"=>"2016", "pdf_views"=>"0", "xml_views"=>"0", "html_views"=>"8"}
  • {"month"=>"8", "year"=>"2016", "pdf_views"=>"5", "xml_views"=>"0", "html_views"=>"29"}
  • {"month"=>"9", "year"=>"2016", "pdf_views"=>"10", "xml_views"=>"0", "html_views"=>"24"}
  • {"month"=>"10", "year"=>"2016", "pdf_views"=>"6", "xml_views"=>"0", "html_views"=>"22"}
  • {"month"=>"11", "year"=>"2016", "pdf_views"=>"2", "xml_views"=>"0", "html_views"=>"17"}
  • {"month"=>"12", "year"=>"2016", "pdf_views"=>"4", "xml_views"=>"0", "html_views"=>"9"}
  • {"month"=>"1", "year"=>"2017", "pdf_views"=>"6", "xml_views"=>"0", "html_views"=>"19"}
  • {"month"=>"2", "year"=>"2017", "pdf_views"=>"2", "xml_views"=>"0", "html_views"=>"23"}
  • {"month"=>"3", "year"=>"2017", "pdf_views"=>"3", "xml_views"=>"0", "html_views"=>"16"}
  • {"month"=>"4", "year"=>"2017", "pdf_views"=>"5", "xml_views"=>"0", "html_views"=>"18"}
  • {"month"=>"5", "year"=>"2017", "pdf_views"=>"5", "xml_views"=>"0", "html_views"=>"18"}
  • {"month"=>"6", "year"=>"2017", "pdf_views"=>"4", "xml_views"=>"0", "html_views"=>"32"}
  • {"month"=>"7", "year"=>"2017", "pdf_views"=>"7", "xml_views"=>"0", "html_views"=>"25"}
  • {"month"=>"8", "year"=>"2017", "pdf_views"=>"6", "xml_views"=>"1", "html_views"=>"16"}
  • {"month"=>"9", "year"=>"2017", "pdf_views"=>"4", "xml_views"=>"0", "html_views"=>"13"}
  • {"month"=>"10", "year"=>"2017", "pdf_views"=>"0", "xml_views"=>"0", "html_views"=>"16"}
  • {"month"=>"11", "year"=>"2017", "pdf_views"=>"3", "xml_views"=>"2", "html_views"=>"20"}
  • {"month"=>"12", "year"=>"2017", "pdf_views"=>"3", "xml_views"=>"0", "html_views"=>"17"}
  • {"month"=>"1", "year"=>"2018", "pdf_views"=>"4", "xml_views"=>"0", "html_views"=>"11"}
  • {"month"=>"2", "year"=>"2018", "pdf_views"=>"4", "xml_views"=>"0", "html_views"=>"5"}
  • {"month"=>"3", "year"=>"2018", "pdf_views"=>"8", "xml_views"=>"3", "html_views"=>"13"}
  • {"month"=>"4", "year"=>"2018", "pdf_views"=>"2", "xml_views"=>"0", "html_views"=>"7"}
  • {"month"=>"5", "year"=>"2018", "pdf_views"=>"6", "xml_views"=>"0", "html_views"=>"10"}
  • {"month"=>"6", "year"=>"2018", "pdf_views"=>"4", "xml_views"=>"0", "html_views"=>"6"}
  • {"month"=>"7", "year"=>"2018", "pdf_views"=>"0", "xml_views"=>"3", "html_views"=>"6"}
  • {"month"=>"8", "year"=>"2018", "pdf_views"=>"5", "xml_views"=>"1", "html_views"=>"9"}
  • {"month"=>"9", "year"=>"2018", "pdf_views"=>"6", "xml_views"=>"0", "html_views"=>"4"}
  • {"month"=>"10", "year"=>"2018", "pdf_views"=>"5", "xml_views"=>"0", "html_views"=>"13"}
  • {"month"=>"11", "year"=>"2018", "pdf_views"=>"2", "xml_views"=>"0", "html_views"=>"12"}
  • {"month"=>"12", "year"=>"2018", "pdf_views"=>"2", "xml_views"=>"0", "html_views"=>"9"}
  • {"month"=>"1", "year"=>"2019", "pdf_views"=>"2", "xml_views"=>"0", "html_views"=>"6"}
  • {"month"=>"2", "year"=>"2019", "pdf_views"=>"5", "xml_views"=>"0", "html_views"=>"13"}
  • {"month"=>"3", "year"=>"2019", "pdf_views"=>"5", "xml_views"=>"1", "html_views"=>"10"}
  • {"month"=>"4", "year"=>"2019", "pdf_views"=>"6", "xml_views"=>"0", "html_views"=>"3"}
  • {"month"=>"5", "year"=>"2019", "pdf_views"=>"9", "xml_views"=>"0", "html_views"=>"18"}
  • {"month"=>"6", "year"=>"2019", "pdf_views"=>"5", "xml_views"=>"0", "html_views"=>"5"}
  • {"month"=>"7", "year"=>"2019", "pdf_views"=>"3", "xml_views"=>"0", "html_views"=>"4"}
  • {"month"=>"8", "year"=>"2019", "pdf_views"=>"4", "xml_views"=>"0", "html_views"=>"13"}
  • {"month"=>"9", "year"=>"2019", "pdf_views"=>"4", "xml_views"=>"0", "html_views"=>"5"}
  • {"month"=>"10", "year"=>"2019", "pdf_views"=>"3", "xml_views"=>"2", "html_views"=>"17"}
  • {"month"=>"11", "year"=>"2019", "pdf_views"=>"2", "xml_views"=>"0", "html_views"=>"1"}
  • {"month"=>"12", "year"=>"2019", "pdf_views"=>"5", "xml_views"=>"0", "html_views"=>"2"}
  • {"month"=>"1", "year"=>"2020", "pdf_views"=>"11", "xml_views"=>"0", "html_views"=>"2"}
  • {"month"=>"2", "year"=>"2020", "pdf_views"=>"3", "xml_views"=>"1", "html_views"=>"4"}

Figshare

  • {"files"=>["https://ndownloader.figshare.com/files/702248"], "description"=>"<p>Western blot analysis of culture supernatants from transgenic <i>P. falciparum</i> clones 3D7_AMA_C_E9 (control) and 3D7_AMA_R_D4 (Ala550Tyr), following schizont rupture in the presence or absence of 10 mM EGTA. Samples were electrophoresed under reducing conditions and probed with anti-PfAMA1 polyclonal serum R5 (top) or monoclonal antibody 24C6.1F1 specific for the parasitophorous vacuole protein SERA5, which is released in the form of the P50 fragment (bottom – loading control). For both clones, shedding of PfAMA1 by PfSUB2 gives rise to the expected products of PfAMA1<sub>48</sub> plus PfAMA1<sub>44</sub> (<a href=\"http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002448#ppat.1002448-Howell2\" target=\"_blank\">[12]</a> and <a href=\"http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002448#ppat-1002448-g001\" target=\"_blank\">Figure 1</a>). In addition, PfAMA1 is shed from the surface of 3D7_AMA_C_E9 parasites via intra-membrane cleavage, giving rise to PfAMA1<sub>52</sub>. This species is absent from 3D7_AMA_R_D4 culture supernatants, confirming that the introduced Ala550Tyr substitution blocks intramembrane processing of PfAMA1. Because PfSUB2-mediated shedding is calcium-dependent, virtually all PfAMA1 shedding is ablated from the 3D7_AMA_R_D4 parasites in the presence of EGTA.</p>", "links"=>[], "tags"=>["intramembrane", "cleavage", "pfama1", "inhibits", "shedding"], "article_id"=>372615, "categories"=>["Biochemistry", "Microbiology"], "users"=>["Anna Olivieri", "Christine R. Collins", "Fiona Hackett", "Chrislaine Withers-Martinez", "Joshua Marshall", "Helen R. Flynn", "J. Mark Skehel", "Michael J. Blackman"], "doi"=>"https://dx.doi.org/10.1371/journal.ppat.1002448.g002", "stats"=>{"downloads"=>0, "page_views"=>0, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Mutation_of_the_intramembrane_cleavage_site_in_PfAMA1_inhibits_shedding_by_intramembrane_processing_/372615", "title"=>"Mutation of the intramembrane cleavage site in PfAMA1 inhibits shedding by intramembrane processing.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2011-12-15 00:43:35"}
  • {"files"=>["https://ndownloader.figshare.com/files/702583"], "description"=>"<p>(A) Schematic representing integration construct intAMA_C (and mutant forms intAMA_P1 and intAMA_P1-P1′) designed to modify the <i>pfama1</i> gene by single-crossover homologous recombination. The wild-type <i>pfama1</i> locus (wt, top) and the modified locus arising from the expected integration even (int) are also shown. <i>Xba</i>I sites (X) used for Southern blot analysis, and the relative positions of oligonucleotide primers used for diagnostic PCR are also indicated (a, primer verAMAint_F; b, primer verAMAint_R – see Table S1 in <a href=\"http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002448#ppat.1002448.s001\" target=\"_blank\">Text S1</a>). Authentic 3D7 <i>pfama1</i> coding sequence is shown in pink, whilst recodonised sequence encoding the FVO <i>pfama1</i> coding sequence is in light blue. The position of the PfSUB2 cleavage site is indicated (black lollypop symbol). p, position of probe used for Southern blot analysis. (B) PCR analysis of genomic DNA from wild-type parent 3D7 and transgenic parasite lines harbouring constructs intAMA_C, intAMA_P1 and intAMA_P1-P1' using primers verAMAint_F and verAMAint_R, schematically represented in panel A. (C) Southern blot analysis of <i>Xba</i>I-digested genomic DNA from wild-type 3D7 and transgenic lines harbouring constructs intAMA_C, intAMA_P1 and intAMA_P1-P1'. The expected sizes of fragments hybridizing with the probe were 12.2 kb for the unmodified wild type locus and 7.0 kb in the case of the expected integration event.</p>", "links"=>[], "tags"=>["shedding", "endogenous", "pfama1", "asexual", "blood-stage", "parasite"], "article_id"=>372949, "categories"=>["Biochemistry", "Microbiology"], "users"=>["Anna Olivieri", "Christine R. Collins", "Fiona Hackett", "Chrislaine Withers-Martinez", "Joshua Marshall", "Helen R. Flynn", "J. Mark Skehel", "Michael J. Blackman"], "doi"=>"https://dx.doi.org/10.1371/journal.ppat.1002448.g006", "stats"=>{"downloads"=>0, "page_views"=>0, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_PfSUB2_mediated_shedding_of_endogenous_PfAMA1_is_essential_for_asexual_blood_stage_parasite_viability_/372949", "title"=>"PfSUB2-mediated shedding of endogenous PfAMA1 is essential for asexual blood-stage parasite viability.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2011-12-15 00:49:09"}
  • {"files"=>["https://ndownloader.figshare.com/files/702477"], "description"=>"<p>(A) and (B) Western blot of schizont extracts and culture supernatants of parental 3D7 and transgenic parasite lines harbouring constructs AMA_C (included in both panels A and B as a control), AMA_R1, AMA_Y1, AMA_Y1-Y1', AMA_F2, AMA_P1, AMA_P1-P1′, and AMA_del. The transgene-derived PfAMA1<sub>83</sub> and PfAMA1<sub>66</sub> products in schizont extracts are detected by anti-HA mAb 3F10. An antibody reactive with the parasitophorous vacuole membrane protein Exp1 serves as a loading control. Culture supernatants collected from the above lines were also probed with the anti-HA mAb 3F10. An antibody (mAb X509) reactive with a shed fragment of MSP1 (MSP1<sub>33</sub>) serves as a loading control. Note that the shed transgenic PfAMA1<sub>48</sub> fragment is virtually absent from supernatants of the AMA_P1, AMA_P1-P1′ and AMA_del mutants. (C) IFA of newly-invaded ring stages of the parasite line harbouring construct AMA_P1. Note that, in contrast to the AMA_C line (not shown), many of the rings detectable with the DNA-reactive dye 4,6-diamidino-2-phenylindole (DAPI; blue) demonstrated strong reactivity with the anti-HA mAb 3F10 (red). The Giemsa-stained field is from a different, stained thin film from the same ring preparation. (D) Quantification of the proportion of anti-HA reactive rings in the AMA_P1, AMA_P1-P1′ and AMA_C lines. The error bars represent standard deviation (SD) values calculated from three different counts of the same experiment.</p>", "links"=>[], "tags"=>["mutagenesis", "cleavage", "prevents", "pfsub2-mediated", "shedding"], "article_id"=>372838, "categories"=>["Biochemistry", "Microbiology"], "users"=>["Anna Olivieri", "Christine R. Collins", "Fiona Hackett", "Chrislaine Withers-Martinez", "Joshua Marshall", "Helen R. Flynn", "J. Mark Skehel", "Michael J. Blackman"], "doi"=>"https://dx.doi.org/10.1371/journal.ppat.1002448.g005", "stats"=>{"downloads"=>0, "page_views"=>0, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Radical_mutagenesis_of_the_cleavage_site_prevents_PfSUB2_mediated_shedding_of_PfAMA1_/372838", "title"=>"Radical mutagenesis of the cleavage site prevents PfSUB2-mediated shedding of PfAMA1.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2011-12-15 00:47:18"}
  • {"files"=>["https://ndownloader.figshare.com/files/702142"], "description"=>"<p>The overall architecture of PfAMA1/DIII-HA is shown schematically (left; not to scale), including the three domains making up the PfAMA1 ectodomain (numbered), the position of the HA epitope tag within the membrane-proximal domain III, the transmembrane domain (TMD; shown as a cylinder), and short cytoplasmic domain. The position of PfSUB2 cleavage, which lies 29 residues from the TMD between Thr517 and Ser518 <a href=\"http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002448#ppat.1002448-Howell2\" target=\"_blank\">[12]</a>, is indicated with a red dot. The nomenclature used for shed fragments of the ectodomain resulting from juxtamembrane or intramembrane cleavage are indicated on the left. Note that PfAMA1<sub>44</sub> is derived from PfAMA1<sub>48</sub> as a result of an internal 'nick' within domain III; both are a result of shedding at the PfSUB2 site, whereas PfAMA1<sub>52</sub> arises from intramembrane shedding <a href=\"http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002448#ppat.1002448-Howell2\" target=\"_blank\">[12]</a>. On the right are shown the names of the parental construct (called AMA_C) for episomal expression of unmodified PfAMA1/DIII-HA, plus the various mutant constructs, together with their corresponding sequence between the PfSUB2 cleavage site (arrowed, with flanking residues in bold) and the TMD (shaded), which contains the intramembrane cleavage site (arrowed). Substitutions and insertions made are highlighted in yellow, whilst residues removed in mutant AMA_del are indicated as dashes. The arrow against the AMA_ins sequence shows the predicted position of PfSUB2 cleavage in this sequence if it is determined primarily by its distance from the TMD. The Ala550Tyr mutation was not expressed episomally, but introduced into the genomic <i>pfama1</i> gene as described in the text. The rat anti-PfAMA1 monoclonal antibody 4G2 used in this work recognizes an epitope formed by domains I and II of PfAMA1 <a href=\"http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002448#ppat.1002448-Collins1\" target=\"_blank\">[32]</a>, <a href=\"http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002448#ppat.1002448-Collins2\" target=\"_blank\">[63]</a>, whilst the rabbit and mouse polyclonal anti-PfAMA1 antibodies used were raised against a recombinant, correctly-folded form of the entire ectodomain, expressed in <i>Pichia pastoris </i><a href=\"http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002448#ppat.1002448-Collins1\" target=\"_blank\">[32]</a>, <a href=\"http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002448#ppat.1002448-Pizarro1\" target=\"_blank\">[64]</a>.</p>", "links"=>[], "tags"=>["antibodies"], "article_id"=>372515, "categories"=>["Biochemistry", "Microbiology"], "users"=>["Anna Olivieri", "Christine R. Collins", "Fiona Hackett", "Chrislaine Withers-Martinez", "Joshua Marshall", "Helen R. Flynn", "J. Mark Skehel", "Michael J. Blackman"], "doi"=>"https://dx.doi.org/10.1371/journal.ppat.1002448.g001", "stats"=>{"downloads"=>1, "page_views"=>5, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Mutants_and_antibodies_used_in_this_study_/372515", "title"=>"Mutants and antibodies used in this study.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2011-12-15 00:41:55"}
  • {"files"=>["https://ndownloader.figshare.com/files/702397"], "description"=>"<p>(A) IFA of drug-selected 3D7 parasites harbouring the AMA_ins construct. The transgene product detected by anti-HA mAb 3F10 (green) co-localises correctly with the microneme protein EBA-175 (red) in mature segmented schizonts. (B) Western blot of parental 3D7 and transgenic parasite lines harbouring constructs AMA_C and AMA_ins. The transgene-derived PfAMA1<sub>83</sub> and PfAMA1<sub>66</sub> products are detected by anti-HA mAb 3F10. Note that both species migrate more slowly in the case of AMA_ins, consistent with the 21-residue insertion in this construct. An antibody reactive with the parasite membrane protein Exp1 is used as a loading control. (C) Western blot of culture supernatants collected from the above lines. The shed transgenic PfAMA1<sub>48</sub> fragment (detected with anti-HA mAb 3F10) is clearly of higher molecular mass in the case of AMA_ins than in the case of the 'wild-type' AMA_C construct. A mAb (X509) reactive with a shed fragment of MSP1 (MSP1<sub>33</sub>) serves as a loading control.</p>", "links"=>[], "tags"=>["pfsub2", "cleavage", "pfama1", "depends"], "article_id"=>372765, "categories"=>["Biochemistry", "Microbiology"], "users"=>["Anna Olivieri", "Christine R. Collins", "Fiona Hackett", "Chrislaine Withers-Martinez", "Joshua Marshall", "Helen R. Flynn", "J. Mark Skehel", "Michael J. Blackman"], "doi"=>"https://dx.doi.org/10.1371/journal.ppat.1002448.g004", "stats"=>{"downloads"=>0, "page_views"=>0, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_The_site_of_PfSUB2_cleavage_in_PfAMA1_depends_primarily_upon_its_distance_from_the_TMD_/372765", "title"=>"The site of PfSUB2 cleavage in PfAMA1 depends primarily upon its distance from the TMD.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2011-12-15 00:46:05"}
  • {"files"=>["https://ndownloader.figshare.com/files/356659"], "description"=>"<div><p>The malarial life cycle involves repeated rounds of intraerythrocytic replication interspersed by host cell rupture which releases merozoites that rapidly invade fresh erythrocytes. Apical membrane antigen-1 (AMA1) is a merozoite protein that plays a critical role in invasion. Antibodies against AMA1 prevent invasion and can protect against malaria <em>in vivo</em>, so AMA1 is of interest as a malaria vaccine candidate. AMA1 is efficiently shed from the invading parasite surface, predominantly through juxtamembrane cleavage by a membrane-bound protease called SUB2, but also by limited intramembrane cleavage. We have investigated the structural requirements for shedding of <em>Plasmodium falciparum</em> AMA1 (PfAMA1), and the consequences of its inhibition. Mutagenesis of the intramembrane cleavage site by targeted homologous recombination abolished intramembrane cleavage with no effect on parasite viability <em>in vitro</em>. Examination of PfSUB2-mediated shedding of episomally-expressed PfAMA1 revealed that the position of cleavage is determined primarily by its distance from the parasite membrane. Certain mutations at the PfSUB2 cleavage site block shedding, and parasites expressing these non-cleavable forms of PfAMA1 on a background of expression of the wild type gene invade and replicate normally <em>in vitro</em>. The non-cleavable PfAMA1 is also functional in invasion. However – in contrast to the intramembrane cleavage site - mutations that block PfSUB2-mediated shedding could not be stably introduced into the genomic <em>pfama1</em> locus, indicating that some shedding of PfAMA1 by PfSUB2 is essential. Remarkably, parasites expressing shedding-resistant forms of PfAMA1 exhibit enhanced sensitivity to antibody-mediated inhibition of invasion. Drugs that inhibit PfSUB2 activity should block parasite replication and may also enhance the efficacy of vaccines based on AMA1 and other merozoite surface proteins.</p> </div>", "links"=>[], "tags"=>["juxtamembrane", "shedding", "ama1", "helps", "evade", "invasion-inhibitory", "antibodies"], "article_id"=>130474, "categories"=>["Biochemistry", "Microbiology"], "users"=>["Anna Olivieri", "Christine R. Collins", "Fiona Hackett", "Chrislaine Withers-Martinez", "Joshua Marshall", "Helen R. Flynn", "J. Mark Skehel", "Michael J. Blackman"], "doi"=>"https://dx.doi.org/10.1371/journal.ppat.1002448", "stats"=>{"downloads"=>12, "page_views"=>7, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/Juxtamembrane_Shedding_of_Plasmodium_falciparum_AMA1_Is_Sequence_Independent_and_Essential_and_Helps_Evade_Invasion_Inhibitory_Antibodies/130474", "title"=>"Juxtamembrane Shedding of <em>Plasmodium falciparum</em> AMA1 Is Sequence Independent and Essential, and Helps Evade Invasion-Inhibitory Antibodies", "pos_in_sequence"=>0, "defined_type"=>3, "published_date"=>"2011-12-15 00:07:54"}
  • {"files"=>["https://ndownloader.figshare.com/files/702648"], "description"=>"<p>Histogram showing the effects of peptide R1 (100 µg ml<sup>−1</sup> final concentration) on erythrocyte invasion by wild type 3D7 parasites or transgenic lines harbouring constructs AMA_C, AMA_P1 or AMA_P1-P1′. Normalised relative invasion efficiencies (expressed as % of controls) were calculated by dividing the parasitaemia values of the R1-treated cultures with those of parallel cultures allowed to undergo invasion in the absence of R1. Error bars indicate standard deviation values, calculated from 5 independent experiments.</p>", "links"=>[], "tags"=>["complementation", "episomally-expressed", "pfama1", "unaffected", "mutations", "pfsub2-mediated"], "article_id"=>373013, "categories"=>["Biochemistry", "Microbiology"], "users"=>["Anna Olivieri", "Christine R. Collins", "Fiona Hackett", "Chrislaine Withers-Martinez", "Joshua Marshall", "Helen R. Flynn", "J. Mark Skehel", "Michael J. Blackman"], "doi"=>"https://dx.doi.org/10.1371/journal.ppat.1002448.g007", "stats"=>{"downloads"=>0, "page_views"=>0, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Functional_complementation_by_episomally_expressed_PfAMA1_is_unaffected_by_mutations_that_block_PfSUB2_mediated_shedding_/373013", "title"=>"Functional complementation by episomally-expressed PfAMA1 is unaffected by mutations that block PfSUB2-mediated shedding.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2011-12-15 00:50:13"}
  • {"files"=>["https://ndownloader.figshare.com/files/702728"], "description"=>"<p>Histogram showing the effects of anti-PfAMA1 antibodies on invasion by transgenic <i>P. falciparum</i> lines harbouring PfAMA1/DIII-HA expression constructs AMA_C, AMA_P1 or AMA_P1-P1′. All assays were performed in the presence of peptide R1 (100 µg ml<sup>−1</sup> final concentration) to block the function of endogenous PfAMA1. Antibodies were used at 0.2 mg ml<sup>−1</sup>. Normalised relative invasion efficiencies (expressed as %) were calculated by dividing the parasitaemia values of the antibody-treated cultures with those of parallel cultures allowed to undergo invasion in the absence of antibodies (peptide R1 only). Error bars indicate standard deviation values.</p>", "links"=>[], "tags"=>["pfsub2-mediated", "shedding", "pfama1", "invasion-inhibitory", "anti-pfama1"], "article_id"=>373092, "categories"=>["Biochemistry", "Microbiology"], "users"=>["Anna Olivieri", "Christine R. Collins", "Fiona Hackett", "Chrislaine Withers-Martinez", "Joshua Marshall", "Helen R. Flynn", "J. Mark Skehel", "Michael J. Blackman"], "doi"=>"https://dx.doi.org/10.1371/journal.ppat.1002448.g008", "stats"=>{"downloads"=>0, "page_views"=>5, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Mutations_that_prevent_PfSUB2_mediated_shedding_of_PfAMA1_increase_sensitivity_to_invasion_inhibitory_anti_PfAMA1_antibodies_/373092", "title"=>"Mutations that prevent PfSUB2-mediated shedding of PfAMA1 increase sensitivity to invasion-inhibitory anti-PfAMA1 antibodies.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2011-12-15 00:51:32"}
  • {"files"=>["https://ndownloader.figshare.com/files/702336"], "description"=>"<p>(A) IFA of drug-selected 3D7 parasites harbouring the AMA_M3 construct. The transgenic PfAMA1/DIII-HA product detected by anti-HA mAb 3F10 (green) co-localises with the microneme protein EBA-175 (red) in mature segmented schizonts. The inset shows similar staining of a free merozoite, where some translocation of PfAMA1/DIII-HA onto the merozoite surface is detectable as is normal for endogenous PfAMA1. Identical IFA patterns were observed in parasites harbouring mutant constructs AMA_M1 and AMA_M2 (not shown). (B) Western blot of parental 3D7 and transgenic parasite lines harbouring construct AMA_C and mutants thereof. All show the characteristic doublet corresponding to PfAMA1<sub>83</sub> (the full-length protein minus signal peptide) and PfAMA1<sub>66</sub> (PfAMA1<sub>83</sub> minus its prosequence) <a href=\"http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002448#ppat.1002448-Howell2\" target=\"_blank\">[12]</a>, <a href=\"http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002448#ppat.1002448-Howell3\" target=\"_blank\">[66]</a>. The transgene products are detected by anti-HA mAb 3F10 in all lines except the parental 3D7 clone. (C) Western blot of culture supernatants collected from the above lines. The shed PfAMA1<sub>48</sub> fragment derived from the transgenes is detected with anti-HA mAb 3F10, whilst a mAb (X509) reactive with a shed fragment of MSP1 (MSP1<sub>33</sub>) serves as a loading control. Note that the single HA tag in PfAMA1/DIII-HA spans the site of the internal 'nick' in domain III that normally produces PfAMA1<sub>44</sub>, preventing the usual partial cleavage at that site <a href=\"http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1002448#ppat.1002448-Collins1\" target=\"_blank\">[32]</a>; PfAMA1<sub>44</sub> is therefore never observed when probing culture supernatants with the anti-HA antibodies.</p>", "links"=>[], "tags"=>["mutagenesis", "pfsub2", "cleavage", "localisation", "shedding"], "article_id"=>372703, "categories"=>["Biochemistry", "Microbiology"], "users"=>["Anna Olivieri", "Christine R. Collins", "Fiona Hackett", "Chrislaine Withers-Martinez", "Joshua Marshall", "Helen R. Flynn", "J. Mark Skehel", "Michael J. Blackman"], "doi"=>"https://dx.doi.org/10.1371/journal.ppat.1002448.g003", "stats"=>{"downloads"=>0, "page_views"=>2, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Alanine_mutagenesis_of_the_PfSUB2_cleavage_site_does_not_affect_expression_localisation_or_shedding_of_PfAMA1_/372703", "title"=>"Alanine mutagenesis of the PfSUB2 cleavage site does not affect expression, localisation or shedding of PfAMA1.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2011-12-15 00:45:03"}

PMC Usage Stats | Further Information

  • {"month"=>"12", "scanned-page-browse"=>"0", "cited-by"=>"0", "abstract"=>"0", "full-text"=>"23", "year"=>"2011", "pdf"=>"13", "unique-ip"=>"18", "figure"=>"11", "scanned-summary"=>"0", "supp-data"=>"0"}
  • {"scanned-page-browse"=>"0", "month"=>"1", "cited-by"=>"0", "abstract"=>"0", "full-text"=>"36", "unique-ip"=>"33", "pdf"=>"32", "year"=>"2012", "figure"=>"7", "scanned-summary"=>"0", "supp-data"=>"4"}
  • {"month"=>"2", "scanned-page-browse"=>"0", "cited-by"=>"0", "abstract"=>"0", "full-text"=>"44", "year"=>"2012", "pdf"=>"30", "unique-ip"=>"36", "figure"=>"11", "scanned-summary"=>"0", "supp-data"=>"1"}
  • {"scanned-page-browse"=>"0", "month"=>"3", "cited-by"=>"0", "abstract"=>"2", "full-text"=>"30", "unique-ip"=>"27", "pdf"=>"19", "year"=>"2012", "figure"=>"5", "scanned-summary"=>"0", "supp-data"=>"0"}
  • {"month"=>"4", "scanned-page-browse"=>"0", "cited-by"=>"0", "abstract"=>"1", "full-text"=>"23", "year"=>"2012", "pdf"=>"12", "unique-ip"=>"23", "figure"=>"17", "scanned-summary"=>"0", "supp-data"=>"1"}
  • {"scanned-page-browse"=>"0", "month"=>"5", "cited-by"=>"0", "abstract"=>"2", "full-text"=>"17", "unique-ip"=>"18", "pdf"=>"9", "year"=>"2012", "figure"=>"5", "scanned-summary"=>"0", "supp-data"=>"0"}
  • {"month"=>"6", "scanned-page-browse"=>"0", "cited-by"=>"0", "abstract"=>"0", "full-text"=>"6", "year"=>"2012", "pdf"=>"5", "unique-ip"=>"7", "figure"=>"1", "scanned-summary"=>"0", "supp-data"=>"0"}
  • {"unique-ip"=>"14", "full-text"=>"13", "pdf"=>"11", "abstract"=>"1", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"3", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2012", "month"=>"7"}
  • {"unique-ip"=>"13", "full-text"=>"13", "pdf"=>"13", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"3", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2012", "month"=>"8"}
  • {"unique-ip"=>"15", "full-text"=>"12", "pdf"=>"6", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"3", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2012", "month"=>"9"}
  • {"unique-ip"=>"14", "full-text"=>"13", "pdf"=>"9", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"3", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2012", "month"=>"10"}
  • {"unique-ip"=>"5", "full-text"=>"4", "pdf"=>"2", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"2", "supp-data"=>"1", "cited-by"=>"0", "year"=>"2012", "month"=>"12"}
  • {"unique-ip"=>"7", "full-text"=>"6", "pdf"=>"3", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"1", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2013", "month"=>"1"}
  • {"unique-ip"=>"8", "full-text"=>"7", "pdf"=>"0", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"2", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2013", "month"=>"2"}
  • {"unique-ip"=>"13", "full-text"=>"9", "pdf"=>"1", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"9", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2013", "month"=>"3"}
  • {"unique-ip"=>"4", "full-text"=>"3", "pdf"=>"1", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"1", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2013", "month"=>"4"}
  • {"unique-ip"=>"7", "full-text"=>"7", "pdf"=>"3", "abstract"=>"1", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"2", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2012", "month"=>"11"}
  • {"unique-ip"=>"11", "full-text"=>"12", "pdf"=>"7", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"13", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2013", "month"=>"5"}
  • {"unique-ip"=>"12", "full-text"=>"10", "pdf"=>"7", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2013", "month"=>"6"}
  • {"unique-ip"=>"5", "full-text"=>"4", "pdf"=>"5", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"2", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2013", "month"=>"7"}
  • {"unique-ip"=>"5", "full-text"=>"2", "pdf"=>"0", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"6", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2013", "month"=>"8"}
  • {"unique-ip"=>"7", "full-text"=>"9", "pdf"=>"1", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2013", "month"=>"9"}
  • {"unique-ip"=>"2", "full-text"=>"1", "pdf"=>"0", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"2", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2013", "month"=>"10"}
  • {"unique-ip"=>"4", "full-text"=>"5", "pdf"=>"1", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2013", "month"=>"11"}
  • {"unique-ip"=>"1", "full-text"=>"1", "pdf"=>"0", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2013", "month"=>"12"}
  • {"unique-ip"=>"10", "full-text"=>"8", "pdf"=>"6", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"2", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2014", "month"=>"1"}
  • {"unique-ip"=>"4", "full-text"=>"4", "pdf"=>"2", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2014", "month"=>"2"}
  • {"unique-ip"=>"9", "full-text"=>"10", "pdf"=>"2", "abstract"=>"1", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2014", "month"=>"3"}
  • {"unique-ip"=>"13", "full-text"=>"11", "pdf"=>"19", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"1", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2014", "month"=>"5"}
  • {"unique-ip"=>"12", "full-text"=>"9", "pdf"=>"3", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"8", "supp-data"=>"1", "cited-by"=>"0", "year"=>"2014", "month"=>"6"}
  • {"unique-ip"=>"8", "full-text"=>"6", "pdf"=>"4", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"1", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2014", "month"=>"4"}
  • {"unique-ip"=>"10", "full-text"=>"5", "pdf"=>"2", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"10", "supp-data"=>"1", "cited-by"=>"0", "year"=>"2014", "month"=>"7"}
  • {"unique-ip"=>"5", "full-text"=>"1", "pdf"=>"4", "abstract"=>"1", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2014", "month"=>"8"}
  • {"unique-ip"=>"12", "full-text"=>"7", "pdf"=>"3", "abstract"=>"2", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"2", "supp-data"=>"1", "cited-by"=>"0", "year"=>"2014", "month"=>"9"}
  • {"unique-ip"=>"11", "full-text"=>"6", "pdf"=>"1", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"4", "supp-data"=>"1", "cited-by"=>"0", "year"=>"2014", "month"=>"10"}
  • {"unique-ip"=>"11", "full-text"=>"10", "pdf"=>"2", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"1", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2014", "month"=>"11"}
  • {"unique-ip"=>"5", "full-text"=>"4", "pdf"=>"0", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"1", "supp-data"=>"1", "cited-by"=>"0", "year"=>"2014", "month"=>"12"}
  • {"unique-ip"=>"6", "full-text"=>"4", "pdf"=>"1", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"11", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2015", "month"=>"1"}
  • {"unique-ip"=>"2", "full-text"=>"2", "pdf"=>"2", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2015", "month"=>"2"}
  • {"unique-ip"=>"6", "full-text"=>"5", "pdf"=>"0", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"1", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2015", "month"=>"3"}
  • {"unique-ip"=>"5", "full-text"=>"5", "pdf"=>"1", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"5", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2015", "month"=>"4"}
  • {"unique-ip"=>"8", "full-text"=>"6", "pdf"=>"1", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"5", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2015", "month"=>"5"}
  • {"unique-ip"=>"7", "full-text"=>"6", "pdf"=>"0", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"5", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2015", "month"=>"6"}
  • {"unique-ip"=>"11", "full-text"=>"12", "pdf"=>"2", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"4", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2015", "month"=>"7"}
  • {"unique-ip"=>"12", "full-text"=>"13", "pdf"=>"2", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"3", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2015", "month"=>"8"}
  • {"unique-ip"=>"4", "full-text"=>"4", "pdf"=>"0", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"1", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2015", "month"=>"9"}
  • {"unique-ip"=>"9", "full-text"=>"12", "pdf"=>"4", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"4", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2015", "month"=>"10"}
  • {"unique-ip"=>"10", "full-text"=>"5", "pdf"=>"4", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"7", "supp-data"=>"0", "cited-by"=>"1", "year"=>"2015", "month"=>"11"}
  • {"unique-ip"=>"7", "full-text"=>"5", "pdf"=>"3", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"1", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2015", "month"=>"12"}
  • {"unique-ip"=>"2", "full-text"=>"1", "pdf"=>"0", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"2", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2016", "month"=>"1"}
  • {"unique-ip"=>"6", "full-text"=>"11", "pdf"=>"1", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"4", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2016", "month"=>"2"}
  • {"unique-ip"=>"5", "full-text"=>"4", "pdf"=>"0", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"6", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2016", "month"=>"3"}
  • {"unique-ip"=>"9", "full-text"=>"8", "pdf"=>"2", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"2", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2016", "month"=>"4"}
  • {"unique-ip"=>"2", "full-text"=>"2", "pdf"=>"0", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2016", "month"=>"5"}
  • {"unique-ip"=>"5", "full-text"=>"5", "pdf"=>"1", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2016", "month"=>"6"}
  • {"unique-ip"=>"7", "full-text"=>"7", "pdf"=>"3", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2016", "month"=>"7"}
  • {"unique-ip"=>"8", "full-text"=>"10", "pdf"=>"2", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2016", "month"=>"8"}
  • {"unique-ip"=>"6", "full-text"=>"5", "pdf"=>"0", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"2", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2016", "month"=>"9"}
  • {"unique-ip"=>"8", "full-text"=>"7", "pdf"=>"2", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"3", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2016", "month"=>"10"}
  • {"unique-ip"=>"4", "full-text"=>"4", "pdf"=>"1", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2016", "month"=>"11"}
  • {"unique-ip"=>"5", "full-text"=>"8", "pdf"=>"1", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2016", "month"=>"12"}
  • {"unique-ip"=>"3", "full-text"=>"3", "pdf"=>"2", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"2", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2017", "month"=>"1"}
  • {"unique-ip"=>"3", "full-text"=>"3", "pdf"=>"1", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2017", "month"=>"2"}
  • {"unique-ip"=>"4", "full-text"=>"4", "pdf"=>"2", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"1", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2017", "month"=>"3"}
  • {"unique-ip"=>"3", "full-text"=>"3", "pdf"=>"1", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"1", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2017", "month"=>"4"}
  • {"unique-ip"=>"1", "full-text"=>"0", "pdf"=>"2", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2017", "month"=>"5"}
  • {"unique-ip"=>"3", "full-text"=>"3", "pdf"=>"0", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2017", "month"=>"6"}
  • {"unique-ip"=>"8", "full-text"=>"9", "pdf"=>"4", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2017", "month"=>"7"}
  • {"unique-ip"=>"5", "full-text"=>"3", "pdf"=>"0", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"3", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2017", "month"=>"8"}
  • {"unique-ip"=>"3", "full-text"=>"2", "pdf"=>"0", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"13", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2017", "month"=>"9"}
  • {"unique-ip"=>"6", "full-text"=>"4", "pdf"=>"2", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2017", "month"=>"10"}
  • {"unique-ip"=>"6", "full-text"=>"7", "pdf"=>"1", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"3", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2017", "month"=>"11"}
  • {"unique-ip"=>"2", "full-text"=>"2", "pdf"=>"1", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2017", "month"=>"12"}
  • {"unique-ip"=>"5", "full-text"=>"5", "pdf"=>"2", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2018", "month"=>"1"}
  • {"unique-ip"=>"1", "full-text"=>"1", "pdf"=>"0", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2018", "month"=>"2"}
  • {"unique-ip"=>"6", "full-text"=>"5", "pdf"=>"2", "abstract"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2018", "month"=>"3"}
  • {"unique-ip"=>"9", "full-text"=>"10", "pdf"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2019", "month"=>"1"}
  • {"unique-ip"=>"1", "full-text"=>"1", "pdf"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2018", "month"=>"5"}
  • {"unique-ip"=>"4", "full-text"=>"6", "pdf"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2018", "month"=>"4"}
  • {"unique-ip"=>"8", "full-text"=>"4", "pdf"=>"4", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2018", "month"=>"6"}
  • {"unique-ip"=>"2", "full-text"=>"1", "pdf"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"1", "cited-by"=>"0", "year"=>"2018", "month"=>"7"}
  • {"unique-ip"=>"2", "full-text"=>"4", "pdf"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2018", "month"=>"9"}
  • {"unique-ip"=>"9", "full-text"=>"14", "pdf"=>"3", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2018", "month"=>"10"}
  • {"unique-ip"=>"3", "full-text"=>"3", "pdf"=>"1", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2018", "month"=>"8"}
  • {"unique-ip"=>"6", "full-text"=>"6", "pdf"=>"1", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"1", "cited-by"=>"0", "year"=>"2018", "month"=>"11"}
  • {"unique-ip"=>"6", "full-text"=>"6", "pdf"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"1", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2018", "month"=>"12"}
  • {"unique-ip"=>"5", "full-text"=>"5", "pdf"=>"0", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"1", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2019", "month"=>"2"}
  • {"unique-ip"=>"3", "full-text"=>"3", "pdf"=>"1", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2019", "month"=>"3"}
  • {"unique-ip"=>"3", "full-text"=>"3", "pdf"=>"1", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2019", "month"=>"4"}
  • {"unique-ip"=>"9", "full-text"=>"11", "pdf"=>"2", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2019", "month"=>"5"}
  • {"unique-ip"=>"10", "full-text"=>"9", "pdf"=>"3", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2019", "month"=>"8"}
  • {"unique-ip"=>"6", "full-text"=>"6", "pdf"=>"2", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2019", "month"=>"9"}
  • {"unique-ip"=>"16", "full-text"=>"17", "pdf"=>"1", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2019", "month"=>"10"}
  • {"unique-ip"=>"7", "full-text"=>"4", "pdf"=>"3", "scanned-summary"=>"0", "scanned-page-browse"=>"0", "figure"=>"0", "supp-data"=>"0", "cited-by"=>"0", "year"=>"2019", "month"=>"12"}

Relative Metric

{"start_date"=>"2011-01-01T00:00:00Z", "end_date"=>"2011-12-31T00:00:00Z", "subject_areas"=>[{"subject_area"=>"/Biology and life sciences/Anatomy and physiology", "average_usage"=>[298, 559, 690, 805, 916, 1018, 1115, 1201, 1283, 1356, 1425, 1498, 1565, 1635, 1703, 1767, 1835, 1902, 1962, 2027, 2089, 2150, 2210, 2265]}, {"subject_area"=>"/Biology and life sciences/Organisms", "average_usage"=>[316, 571, 699, 812, 915, 1008, 1095, 1180, 1255, 1319, 1393, 1461, 1528, 1595, 1662, 1729, 1802, 1868, 1937, 1997, 2062, 2130, 2192, 2254, 2324, 2386, 2450, 2510, 2579, 2642, 2716, 2778, 2842, 2908, 2979, 3042, 3095]}, {"subject_area"=>"/Biology and life sciences/Parasitology", "average_usage"=>[359, 649, 766, 882, 961, 1046, 1170, 1246, 1317, 1379, 1458, 1532, 1583, 1643, 1697, 1774, 1867, 1925, 1988, 2025, 2077, 2131, 2221, 2341, 2424, 2471, 2514, 2551, 2613, 2691, 2738, 2792, 2852, 2903, 2979, 3037, 3093]}, {"subject_area"=>"/Biology and life sciences/Physiology", "average_usage"=>[296, 551, 687, 800, 903, 1001, 1095, 1184, 1259, 1327, 1406, 1473, 1535, 1600, 1669, 1736, 1794, 1857, 1922, 1983, 2046, 2109, 2169, 2223, 2278, 2344, 2402, 2457, 2517, 2574, 2621, 2688, 2756, 2811, 2872, 2936, 2985]}, {"subject_area"=>"/Medicine and health sciences/Anatomy and physiology", "average_usage"=>[296, 554, 686, 802, 910, 1014, 1104, 1195, 1277, 1349, 1419, 1483, 1551, 1620, 1690, 1757, 1820, 1888, 1957, 2017, 2081, 2140, 2202, 2259]}]}
Loading … Spinner
There are currently no alerts