NRFL-1, the C. elegans NHERF Orthologue, Interacts with Amino Acid Transporter 6 (AAT-6) for Age-Dependent Maintenance of AAT-6 on the Membrane
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{"title"=>"NRFL-1, the C. elegans NHERF orthologue, interacts with amino acid transporter 6 (AAT-6) for age-dependent maintenance of AAT-6 on the membrane", "type"=>"journal", "authors"=>[{"first_name"=>"Kohei", "last_name"=>"Hagiwara", "scopus_author_id"=>"55337314300"}, {"first_name"=>"Shushi", "last_name"=>"Nagamori", "scopus_author_id"=>"7006761600"}, {"first_name"=>"Yasuhiro M.", "last_name"=>"Umemura", "scopus_author_id"=>"55337519300"}, {"first_name"=>"Ryuichi", "last_name"=>"Ohgaki", "scopus_author_id"=>"8618486100"}, {"first_name"=>"Hidekazu", "last_name"=>"Tanaka", "scopus_author_id"=>"56255034500"}, {"first_name"=>"Daisuke", "last_name"=>"Murata", "scopus_author_id"=>"57062494900"}, {"first_name"=>"Saya", "last_name"=>"Nakagomi", "scopus_author_id"=>"57194481602"}, {"first_name"=>"Kazuko H.", "last_name"=>"Nomura", "scopus_author_id"=>"7402536737"}, {"first_name"=>"Eriko", "last_name"=>"Kage-Nakadai", "scopus_author_id"=>"35078736100"}, {"first_name"=>"Shohei", "last_name"=>"Mitani", "scopus_author_id"=>"56975996700"}, {"first_name"=>"Kazuya", "last_name"=>"Nomura", "scopus_author_id"=>"35102586200"}, {"first_name"=>"Yoshikatsu", "last_name"=>"Kanai", "scopus_author_id"=>"55575124400"}], "year"=>2012, "source"=>"PLoS ONE", "identifiers"=>{"pmid"=>"22916205", "sgr"=>"84865053085", "scopus"=>"2-s2.0-84865053085", "isbn"=>"1932-6203 (Electronic)\\r1932-6203 (Linking)", "doi"=>"10.1371/journal.pone.0043050", "pui"=>"365445656", "issn"=>"19326203"}, "id"=>"1e2382a4-6644-3475-b983-eb8ca7579d2a", "abstract"=>"The NHERF (Na(+)/H(+) exchanger regulatory factor) family has been proposed to play a key role in regulating transmembrane protein localization and retention at the plasma membrane. Due to the high homology between the family members, potential functional compensations have been a concern in sorting out the function of individual NHERF numbers. Here, we studied C. elegans NRFL-1 (C01F6.6) (nherf-like protein 1), the sole C. elegans orthologue of the NHERF family, which makes worm a model with low genetic redundancy of NHERF homologues. Integrating bioinformatic knowledge of C. elegans proteins into yeast two-hybrid scheme, we identified NRFL-1 as an interactor of AAT-6, a member of the C. elegans AAT (amino acid transporter) family. A combination of GST pull-down assay, localization study, and co-immunoprecipitation confirmed the binding and characterized the PDZ interaction. AAT-6 localizes to the luminal membrane even in the absence of NRFL-1 when the worm is up to four-day old. A fluorescence recovery after photobleaching (FRAP) analysis suggested that NRFL-1 immobilizes AAT-6 at the luminal membrane. When the nrfl-1 deficient worm is six-day or older, in contrast, the membranous localization of AAT-6 is not observed, whereas AAT-6 tightly localizes to the membrane in worms with NRFL-1. Sorting out the in vivo functions of the C. elegans NHERF protein, we found that NRFL-1, a PDZ-interactor of AAT-6, is responsible for the immobilization and the age-dependent maintenance of AAT-6 on the intestinal luminal membrane.", "link"=>"http://www.mendeley.com/research/nrfl1-c-elegans-nherf-orthologue-interacts-amino-acid-transporter-6-aat6-agedependent-maintenance-aa", "reader_count"=>5, "reader_count_by_academic_status"=>{"Unspecified"=>1, "Student > Master"=>1, "Student > Bachelor"=>1, "Professor"=>1, "Professor > Associate Professor"=>1}, "reader_count_by_user_role"=>{"Unspecified"=>1, "Student > Master"=>1, "Student > Bachelor"=>1, "Professor"=>1, "Professor > Associate Professor"=>1}, "reader_count_by_subject_area"=>{"Unspecified"=>1, "Agricultural and Biological Sciences"=>4}, "reader_count_by_subdiscipline"=>{"Agricultural and Biological Sciences"=>{"Agricultural and Biological Sciences"=>4}, "Unspecified"=>{"Unspecified"=>1}}, "group_count"=>0}

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Figshare

  • {"files"=>["https://ndownloader.figshare.com/files/311221", "https://ndownloader.figshare.com/files/311279", "https://ndownloader.figshare.com/files/311348", "https://ndownloader.figshare.com/files/311390"], "description"=>"<div><p>The NHERF (Na<sup>+</sup>/H<sup>+</sup> exchanger regulatory factor) family has been proposed to play a key role in regulating transmembrane protein localization and retention at the plasma membrane. Due to the high homology between the family members, potential functional compensations have been a concern in sorting out the function of individual NHERF numbers. Here, we studied <em>C</em>. <em>elegans</em> NRFL-1 (C01F6.6) (nherf-like protein 1), the sole <em>C</em>. <em>elegans</em> orthologue of the NHERF family, which makes worm a model with low genetic redundancy of NHERF homologues. Integrating bioinformatic knowledge of <em>C</em>. <em>elegans</em> proteins into yeast two-hybrid scheme, we identified NRFL-1 as an interactor of AAT-6, a member of the <em>C</em>. <em>elegans</em> AAT (amino acid transporter) family. A combination of GST pull-down assay, localization study, and co-immunoprecipitation confirmed the binding and characterized the PDZ interaction. AAT-6 localizes to the luminal membrane even in the absence of NRFL-1 when the worm is up to four-day old. A fluorescence recovery after photobleaching (FRAP) analysis suggested that NRFL-1 immobilizes AAT-6 at the luminal membrane. When the <em>nrfl-1</em> deficient worm is six-day or older, in contrast, the membranous localization of AAT-6 is not observed, whereas AAT-6 tightly localizes to the membrane in worms with NRFL-1. Sorting out the <em>in vivo</em> functions of the <em>C</em>. <em>elegans</em> NHERF protein, we found that NRFL-1, a PDZ-interactor of AAT-6, is responsible for the immobilization and the age-dependent maintenance of AAT-6 on the intestinal luminal membrane.</p> </div>", "links"=>[], "tags"=>["nherf", "interacts", "amino", "transporter", "age-dependent", "aat-6", "membrane"], "article_id"=>121392, "categories"=>["Biological Sciences", "Biochemistry", "Developmental Biology"], "users"=>["Kohei Hagiwara", "Shushi Nagamori", "Yasuhiro M. Umemura", "Ryuichi Ohgaki", "Hidekazu Tanaka", "Daisuke Murata", "Saya Nakagomi", "Kazuko H. Nomura", "Eriko Kage-Nakadai", "Shohei Mitani", "Kazuya Nomura", "Yoshikatsu Kanai"], "doi"=>["https://dx.doi.org/10.1371/journal.pone.0043050.s001", "https://dx.doi.org/10.1371/journal.pone.0043050.s002", "https://dx.doi.org/10.1371/journal.pone.0043050.s003", "https://dx.doi.org/10.1371/journal.pone.0043050.s004"], "stats"=>{"downloads"=>0, "page_views"=>0, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/NRFL_1_the_C_elegans_NHERF_Orthologue_Interacts_with_Amino_Acid_Transporter_6_AAT_6_for_Age_Dependent_Maintenance_of_AAT_6_on_the_Membrane/121392", "title"=>"NRFL-1, the <em>C</em>. <em>elegans</em> NHERF Orthologue, Interacts with Amino Acid Transporter 6 (AAT-6) for Age-Dependent Maintenance of AAT-6 on the Membrane", "pos_in_sequence"=>0, "defined_type"=>4, "published_date"=>"2012-08-15 00:23:12"}
  • {"files"=>["https://ndownloader.figshare.com/files/591192"], "description"=>"<p><b><i>A</i></b>, Schematic illustration of <i>nrfl-1</i> transcript (modified from WormBase). Black boxes, exons; grey box, spliced leader 1 (SL1); CC, two bases of cytosine constituting the 5′UTR; ATG, the start codon; TAA, the stop codon; AATAAA, a putative polyadenylation signal; poly-A, poly adenosine tail. The alleles <i>tm3501</i> and <i>ok2292</i> lack 953-basepair and 1117-basepair of the genomic DNA, respectively, indicated by the horizontal lines. <b><i>B</i></b>, NRFL-1 protein. The <i>nrfl-1</i>gene products contain either a single or double PDZ domains with PDZ II shared by all. The variants C01F6.6a, c, and e have PDZ I and PDZ II (double-domain group). PDZ I is structurally related to PDZ II with 45% identity and 59% similarity. <b><i>C</i></b>, Pairwise domain comparison between NRFL-1 and two human NHERF proteins: NHERF 1 and NHERF 2. PDZ domains were specified by ExPASy Prosite <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0043050#pone.0043050-Gasteiger1\" target=\"_blank\">[67]</a> and compared by BLAST <a href=\"http://www.plosone.org/article/info:doi/10.1371/journal.pone.0043050#pone.0043050-Altschul1\" target=\"_blank\">[26]</a>. For each pair, identity/similarity (%/%) is assigned. The red lines indicate the most remarkable pair(s) for each of NRFL-1 PDZ domains with respect to sequence identity.</p>", "links"=>[], "tags"=>["worm", "orthologue", "nherf"], "article_id"=>261673, "categories"=>["Biological Sciences", "Biochemistry", "Developmental Biology"], "users"=>["Kohei Hagiwara", "Shushi Nagamori", "Yasuhiro M. Umemura", "Ryuichi Ohgaki", "Hidekazu Tanaka", "Daisuke Murata", "Saya Nakagomi", "Kazuko H. Nomura", "Eriko Kage-Nakadai", "Shohei Mitani", "Kazuya Nomura", "Yoshikatsu Kanai"], "doi"=>["https://dx.doi.org/10.1371/journal.pone.0043050.g001"], "stats"=>{"downloads"=>0, "page_views"=>0, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_NRFL_1_the_worm_orthologue_of_NHERF_family_/261673", "title"=>"NRFL-1, the worm orthologue of NHERF family.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2012-08-15 00:27:53"}
  • {"files"=>["https://ndownloader.figshare.com/files/591292"], "description"=>"<p><b><i>A</i></b>, Yeast-two hybrid assay. Full-length NRFL-1 was used as a prey, whereas the empty bait vector (<i>control</i>), AAT-6 <i>C</i>-terminus tail corresponding to residues 487–523 (<i>AAT-6</i>), and AAT-6 <i>C</i>-terminus tail without PDZ bindings motif (Δ<i>TRM</i>) were used as baits. Each bait-prey pair was assessed for LEU2 and GFP reporters. The pair of AAT-6 <i>C</i>-terminus tail and NRFL-1 grew on the medium without leucine and glowed. Similar results were obtained in three different experiments. <b><i>B</i></b>, GST pull-down assay. 3×FLAG-NRFL-1 was pulled down by GST fusion of AAT-6 <i>C</i>-terminus tail corresponding to residues 487–523 (<i>GST-AAT-6</i>), but not by GST (<i>GST</i>) or GST fusion of AAT-6 <i>C</i>-terminus tail without PDZ binding motif (<i>GST-</i>Δ<i>TRM</i>). <i>Upper</i>, pull-down products probed by anti-FLAG antibody. <i>Lower</i>, pull-down products reprobed by anti-GST antibody. A representative blot from three experiments is shown. <b><i>C</i></b>, Domain analysis by GST pull-down assay. Lanes 1, 2, 3 and 4 are for 3×FLAG-NRFL-1 (wild type), 3×FLAG-NRFL-1 with PDZ I/PDZ II both mutated (G26A/Y27A and E154A/F155A), 3×FLAG-NRFL-1 with PDZ I mutated (G26A/Y27A), and 3×FLAG-NRFL-1 with PDZ II mutated (E154A/F155A), respectively. GST fusion of AAT-6 <i>C</i>-terminus tail corresponding to residues 487–523 (<i>GST-AAT-6</i>) pulled down 3×FLAG-NRFL-1 (wild type) and 3×FLAG-NRFL-1 with PDZ II mutated but not 3×FLAG-NRFL-1 with PDZ I/PDZ II both mutated and 3×FLAG-NRFL-1 with PDZ II mutated, suggesting that PDZ II is the preferred domain for interacting with AAT-6. + and – in the figure denote wild type and mutated domain, respectively. <i>Upper</i>, input (1/10 of sample volume); <i>middle</i>, pull-down products probed by anti-FLAG antibody; <i>bottom</i>, pull-down product reprobed by anti-GST antibody. A representative blot from three experiments is shown.</p>", "links"=>[], "tags"=>["nrfl-1"], "article_id"=>261777, "categories"=>["Biological Sciences", "Biochemistry", "Developmental Biology"], "users"=>["Kohei Hagiwara", "Shushi Nagamori", "Yasuhiro M. Umemura", "Ryuichi Ohgaki", "Hidekazu Tanaka", "Daisuke Murata", "Saya Nakagomi", "Kazuko H. Nomura", "Eriko Kage-Nakadai", "Shohei Mitani", "Kazuya Nomura", "Yoshikatsu Kanai"], "doi"=>["https://dx.doi.org/10.1371/journal.pone.0043050.g002"], "stats"=>{"downloads"=>0, "page_views"=>0, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_PDZ_interaction_between_NRFL_1_and_AAT_6_/261777", "title"=>"PDZ interaction between NRFL-1 and AAT-6.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2012-08-15 00:29:37"}
  • {"files"=>["https://ndownloader.figshare.com/files/591392"], "description"=>"<p><i>–</i><b><i>D</i></b>, Expression of NRFL-1 in worm. <i>gfp::nrfl-1</i> expression was detected in excretory canal (<i>arrow</i> in <b><i>A</i></b> and <b><i>B</i></b>) and luminal membrane of intestinal epithelial cells (<i>single-arrowhead</i> ) in the anterior (<b><i>B</i></b>), middle (<b><i>C</i></b>) and posterior (<b><i>D</i></b>) intestine. <i>gfp::nrfl-1</i> was not detected on the basal side (<i>double-arrowhead</i> in <b><i>B</i></b>, <b><i>C</i></b> and <b><i>D</i></b>). In <b><i>A</i></b>, a worm with <i>gfp::nrfl-1</i> expression restricted to the excretory system was imaged for clarity. Such worms occasionally occurred in the transgenic population. In <b><i>C</i></b> and <b><i>D</i></b>, cytosolic dispersion of NRFL-1 was seen. Scale bars, 25 µm. Ten worms examined for each. <b><i>E</i></b>, Luminal enrichment of NRFL-1. Endogenous NRFL-1 was detected by anti-NRFL-1 antibody along the luminal side of the intestine. Non-specific signal on the body-wall is arrowed (<i>left</i>). IFB-2 was immune-labeled in a similar pattern (<i>middle</i>). Merged image shows that NRFL-1 is distributed apical to IFB-2 (<i>right</i>). Scale bar, 25 µm. Confocal images of a representative intestinal section (whole-worm) from seven independent experiments are shown. <b><i>F</i></b>, Immunoblot of endogenous and recombinant NRFL-1. Left (<i>recombinant</i>), middle (<i>wild type</i>) and right (<i>nrfl-1</i>) lanes are for recombinant C01F6.6a and C01F6.6b proteins (untagged), lysate from wild-type, and lysate from <i>nrfl-1</i>(<i>tm3501</i>), respectively. Note that the band at ∼72 kDa in the middle lane was not detected in the right and the band did not match either C01F6.6a (∼62 kDa) or C01F6.6b (∼78 kDa). Sixty microgram of protein was loaded for the lysates. A representative blot from three separate experiments is shown. <b><i>G</i></b>, Dephosphorylation of endogenous NRFL-1. The wild-type lysate was incubated with (+) and without (−) phosphatase inhibitors for the indicated period. Over incubation, the bands for inhibitor-free lysate migrated towards a position corresponding to the recombinant C01F6.6a. Ninety microgram of protein was loaded for the lysate. The results are confirmed in duplicate experiments.</p>", "links"=>[], "tags"=>["nrfl-1"], "article_id"=>261879, "categories"=>["Biological Sciences", "Biochemistry", "Developmental Biology"], "users"=>["Kohei Hagiwara", "Shushi Nagamori", "Yasuhiro M. Umemura", "Ryuichi Ohgaki", "Hidekazu Tanaka", "Daisuke Murata", "Saya Nakagomi", "Kazuko H. Nomura", "Eriko Kage-Nakadai", "Shohei Mitani", "Kazuya Nomura", "Yoshikatsu Kanai"], "doi"=>["https://dx.doi.org/10.1371/journal.pone.0043050.g003"], "stats"=>{"downloads"=>0, "page_views"=>0, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Expression_and_protein_profile_of_NRFL_1_C01F6_6_in_C_elegans_A_/261879", "title"=>"Expression and protein profile of NRFL-1 (C01F6.6) in <i>C. elegans</i>. <i>A</i>", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2012-08-15 00:31:19"}
  • {"files"=>["https://ndownloader.figshare.com/files/591529"], "description"=>"<p>, Expression of AAT-6 in worms. AAT-6<i><sup>1−517</sup></i>::GFP::AAT-6<i><sup>518–523</sup></i> was localized to the luminal surface (<i>single arrowhead</i>) but not to the basal side (<i>double arrowhead</i>) of the intestinal epithelia. Scale bar, 25 µm. Non-specific fluorescence on gut granules was seen. More than twenty worms were analyzed. <b><i>B</i></b>, Co-localization of AAT-6 and NRFL-1. GFP fluorescence from AAT-6<i><sup>1−517</sup></i>::GFP::AAT-6<i><sup>518–523</sup></i> (<i>top</i>) was co-localized with immunostaining of NRFL-1 by anti-NRFL-1 antibody visualized by Cy3-labeled secondary antibody (<i>middle</i>). Bottom image is merged from top and middle images. Confocal images of a representative intestine section (whole worm) are shown. Scale bar: 25 µm. More than five worms were analyzed. <b><i>C</i></b>, Intensity profile along the line A–B in the merged image shows an overlapping of the peaks of the NRFL-1and the AAT-6 signal. <b><i>D</i></b>, Immunoprecipitation of NRFL-1/AAT-6 complex from worm lysate. The worm lysate was immunoprecipitated with anti-GFP monoclonal antibody (mouse) and immunoblotted using anti-NRFL-1 antibody. <i>Top</i>: input (2.5%). <i>Middle</i> and <i>bottom</i>: immunoprecipitant was immunoblotted using anti-NRFL-1 antibody and anti-GFP antibody (chicken), respectively. In the middle blot, two major bands of NRFL-1 detected (<i>arrowheads</i>), probably reflecting partial dephosphorylation during immunoprecipitation process. A representative blot of two separate experiments is shown.</p>", "links"=>[], "tags"=>["aat-6", "nrfl-1"], "article_id"=>262012, "categories"=>["Biological Sciences", "Biochemistry", "Developmental Biology"], "users"=>["Kohei Hagiwara", "Shushi Nagamori", "Yasuhiro M. Umemura", "Ryuichi Ohgaki", "Hidekazu Tanaka", "Daisuke Murata", "Saya Nakagomi", "Kazuko H. Nomura", "Eriko Kage-Nakadai", "Shohei Mitani", "Kazuya Nomura", "Yoshikatsu Kanai"], "doi"=>["https://dx.doi.org/10.1371/journal.pone.0043050.g004"], "stats"=>{"downloads"=>0, "page_views"=>0, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Interaction_of_AAT_6_with_NRFL_1_in_vivo_A_/262012", "title"=>"Interaction of AAT-6 with NRFL-1 <i>in vivo</i>. <i>A</i>", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2012-08-15 00:33:32"}
  • {"files"=>["https://ndownloader.figshare.com/files/591650"], "description"=>"<p>, The localization of AAT-6<i><sup>1−517</sup></i>::GFP::AAT-6<i><sup>518–523</sup></i> was compared between <i>aat-6</i> and <i>nrfl-1</i>(<i>tm3501</i>);<i>aat-6</i> genetic backgrounds. Epifluorescence images of the distribution of AAT-6 in the intestine are shown for worms two, four and six days after hatching. In six-day old worm, the membranous localization of AAT-6 decayed in <i>nrfl-1</i>(<i>tm3501</i>);<i>aat-6</i>, whereas AAT-6 was retained on the luminal membrane in six days in the presence of NRFL-1 (<i>aat-6</i>). Scale bars: 100 µm. Representative pictures from more than ten worms analyzed for each are shown. <b><i>B</i></b>, Fluorescence intensity was measured to quantify the age-dependent regulation. The intensity of the intestinal luminal surface was peaked at day four with significantly stronger signal in <i>aat-6</i> (gray column) compared with <i>nrfl-1</i>(<i>tm3501</i>);<i>aat-6</i> (white column). The intensity at day six did not differ significantly between the strains (<i>luminal membrane</i>). <b><i>C</i></b>, The fluorescence intensity of the whole intestine showed a similar pattern (<i>whole intestine</i>). The localization index, luminal intensity divided by whole intestine intensity, quantifies the membranous localization. The six-day old <i>nrfl-1</i>(<i>tm3501</i>);<i>aat-6</i> worm had a significantly lower score, showing age-dependent decay in luminal localization (<i>localization index</i>). Gray column, <i>aat-6</i>. White column, <i>nrfl-1</i>(<i>tm3501</i>);<i>aat-6</i>. Values are presented with mean ± S.E. (n = 5). <b><i>D</i></b>, Immunoblot and densitometric analysis of AAT-6<i><sup>1−517</sup></i>::GFP::AAT-6<i><sup>518–523</sup></i> in six-day old worm. Densitometric analysis followed by anti-GFP antibody immunobloting exhibited no significant difference in band intensity between the genetic backgrounds. A representative blot was presented with actin as a loading control. The bar graph indicates the relative band intensities of the respective sample. Values are presented with mean ± S.E. (n = 4).</p>", "links"=>[], "tags"=>["aat-6", "intestinal", "luminal", "membrane"], "article_id"=>262131, "categories"=>["Biological Sciences", "Biochemistry", "Developmental Biology"], "users"=>["Kohei Hagiwara", "Shushi Nagamori", "Yasuhiro M. Umemura", "Ryuichi Ohgaki", "Hidekazu Tanaka", "Daisuke Murata", "Saya Nakagomi", "Kazuko H. Nomura", "Eriko Kage-Nakadai", "Shohei Mitani", "Kazuya Nomura", "Yoshikatsu Kanai"], "doi"=>["https://dx.doi.org/10.1371/journal.pone.0043050.g005"], "stats"=>{"downloads"=>0, "page_views"=>0, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Maintenance_of_AAT_6_on_the_intestinal_luminal_membrane_by_NRFL_1_A_/262131", "title"=>"Maintenance of AAT-6 on the intestinal luminal membrane by NRFL-1. <i>A</i>", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2012-08-15 00:35:31"}
  • {"files"=>["https://ndownloader.figshare.com/files/591813"], "description"=>"<p><b><i>A</i></b>, FRAP analysis of the AAT-6 dynamics was performed in 4-day old worms. Confocal images of AAT-6 (AAT-6<i><sup>1−517</sup></i>::GFP::AAT-6<i><sup>518–523</sup></i>) was compared between <i>aat-6</i> and <i>nrfl-1</i>;<i>aat-6</i> genetic backgrounds. Top pictures indicate representative images prior to photobleaching (<i>pre-bleach</i>), immediately after photobleaching (<i>post-bleach</i>), and 300 sec after photobleaching (<i>recovery</i>). <b><i>B</i></b>, Graph depicts the time course of recovery of AAT-6 fluorescence for <i>nrfl-1</i>;<i>aat-6</i> (•) and <i>aat-6</i> (□). Recovery was measured with the pre-bleach fluorescence intensity being 100% and the post-bleach intensity being 0%. The recovery curves were generated from 5 separate experiments and the values were expressed as mean ± S.E. (n = 5). Scale bars: 2 µm.</p>", "links"=>[], "tags"=>["aat-6", "intestinal", "apical", "membrane"], "article_id"=>262296, "categories"=>["Biological Sciences", "Biochemistry", "Developmental Biology"], "users"=>["Kohei Hagiwara", "Shushi Nagamori", "Yasuhiro M. Umemura", "Ryuichi Ohgaki", "Hidekazu Tanaka", "Daisuke Murata", "Saya Nakagomi", "Kazuko H. Nomura", "Eriko Kage-Nakadai", "Shohei Mitani", "Kazuya Nomura", "Yoshikatsu Kanai"], "doi"=>["https://dx.doi.org/10.1371/journal.pone.0043050.g006"], "stats"=>{"downloads"=>0, "page_views"=>0, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Immobilization_of_AAT_6_on_the_intestinal_apical_membrane_by_NRFL_1_/262296", "title"=>"Immobilization of AAT-6 on the intestinal apical membrane by NRFL-1.", "pos_in_sequence"=>0, "defined_type"=>1, "published_date"=>"2012-08-15 00:38:16"}
  • {"files"=>["https://ndownloader.figshare.com/files/591901"], "description"=>"<p>Each of the sixteen intestinal PDZ proteins (prey) was subjected to a yeast two-hybrid assay with AAT-6 as a bait. Pairs were assessed for LEU2 and GFP reporter genes: + and −, positive and negative for LEU2 or GFP, respectively.</p>", "links"=>[], "tags"=>["two-hybrid"], "article_id"=>262390, "categories"=>["Biological Sciences", "Biochemistry", "Developmental Biology"], "users"=>["Kohei Hagiwara", "Shushi Nagamori", "Yasuhiro M. Umemura", "Ryuichi Ohgaki", "Hidekazu Tanaka", "Daisuke Murata", "Saya Nakagomi", "Kazuko H. Nomura", "Eriko Kage-Nakadai", "Shohei Mitani", "Kazuya Nomura", "Yoshikatsu Kanai"], "doi"=>["https://dx.doi.org/10.1371/journal.pone.0043050.t001"], "stats"=>{"downloads"=>0, "page_views"=>0, "likes"=>0}, "figshare_url"=>"https://figshare.com/articles/_Yeast_two_hybrid_matching_/262390", "title"=>"Yeast two-hybrid matching.", "pos_in_sequence"=>0, "defined_type"=>3, "published_date"=>"2012-08-15 00:39:50"}

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Relative Metric

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