{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,13]],"date-time":"2026-10-13T14:16:37Z","timestamp":1791900997858,"version":"4.3.4"},"reference-count":41,"publisher":"American Association for the Advancement of Science (AAAS)","issue":"5424","content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Science"],"published-print":{"date-parts":[[1999,7,2]]},"abstract":"<jats:p>\n            Epithelia permit selective and regulated flux from apical to basolateral surfaces by transcellular passage through cells or paracellular flux between cells. Tight junctions constitute the barrier to paracellular conductance; however, little is known about the specific molecules that mediate paracellular permeabilities. Renal magnesium ion (Mg\n            <jats:sup>2+<\/jats:sup>\n            ) resorption occurs predominantly through a paracellular conductance in the thick ascending limb of Henle (TAL). Here, positional cloning has identified a human gene,\n            <jats:italic>paracellin-1<\/jats:italic>\n            (\n            <jats:italic>PCLN-1<\/jats:italic>\n            ), mutations in which cause renal Mg\n            <jats:sup>2+<\/jats:sup>\n            wasting. PCLN-1 is located in tight junctions of the TAL and is related to the claudin family of tight junction proteins. These findings provide insight into Mg\n            <jats:sup>2+<\/jats:sup>\n            homeostasis, demonstrate the role of a tight junction protein in human disease, and identify an essential component of a selective paracellular conductance.\n          <\/jats:p>","DOI":"10.1126\/science.285.5424.103","type":"journal-article","created":{"date-parts":[[2002,7,27]],"date-time":"2002-07-27T09:49:42Z","timestamp":1027763382000},"page":"103-106","source":"Crossref","is-referenced-by-count":977,"title":["Paracellin-1, a Renal Tight Junction Protein Required for Paracellular Mg\n            <sup>2+<\/sup>\n            Resorption"],"prefix":"10.1126","volume":"285","author":[{"given":"David B.","family":"Simon","sequence":"first","affiliation":[{"name":"Howard Hughes Medical Institute, Department of Genetics,"},{"name":"Department of Medicine, Yale University School of Medicine, New Haven, CT 06510, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yin","family":"Lu","sequence":"additional","affiliation":[{"name":"Howard Hughes Medical Institute, Department of Genetics,"},{"name":"Department of Medicine, Yale University School of Medicine, New Haven, CT 06510, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Keith A.","family":"Choate","sequence":"additional","affiliation":[{"name":"Howard Hughes Medical Institute, Department of Genetics,"},{"name":"Department of Medicine, Yale University School of Medicine, New Haven, CT 06510, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Heino","family":"Velazquez","sequence":"additional","affiliation":[{"name":"Department of Medicine, Yale University School of Medicine, New Haven, CT 06510, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Essam","family":"Al-Sabban","sequence":"additional","affiliation":[{"name":"Department of Pediatrics, King Faisal Specialist Hospital and Research Center, Riyadh, Saudi Arabia."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Manuel","family":"Praga","sequence":"additional","affiliation":[{"name":"Servicio de Nefrologia, Hospital 12 de Octubre, Madrid, Spain."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Giorgio","family":"Casari","sequence":"additional","affiliation":[{"name":"Telethon Institute of Genetics and Medicine, Milan 20132, Italy."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alberto","family":"Bettinelli","sequence":"additional","affiliation":[{"name":"Department of Pediatrics II, University of Milan, Milan, Italy."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Giacomo","family":"Colussi","sequence":"additional","affiliation":[{"name":"Renal Division, Niguarda \u201cCa' Granda\u201d Hospital, Milan, Italy."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Juan","family":"Rodriguez-Soriano","sequence":"additional","affiliation":[{"name":"Department of Pediatrics, Hospital de Cruces, Baracaldo E-48903, Spain."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"David","family":"McCredie","sequence":"additional","affiliation":[{"name":"Department of Nephrology, Royal Children's Hospital, Flemington Road, Parkville, Victoria 3052, Australia."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"David","family":"Milford","sequence":"additional","affiliation":[{"name":"Department of Nephrology, Birmingham Children's Hospital, Birmingham B16 8ET, UK."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sami","family":"Sanjad","sequence":"additional","affiliation":[{"name":"Department of Pediatrics, American University, Beirut, Lebanon."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Richard P.","family":"Lifton","sequence":"additional","affiliation":[{"name":"Howard Hughes Medical Institute, Department of Genetics,"},{"name":"Department of Medicine, Yale University School of Medicine, New Haven, CT 06510, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"221","reference":[{"key":"e_1_3_1_2_2","first-page":"58","volume":"21","author":"Bonnet R.","year":"1895","unstructured":"Bonnet R., Dtsch. Med. Wochenschr. 21, 58 (1895);","journal-title":"Dtsch. Med. Wochenschr."},{"key":"e_1_3_1_2_3","doi-asserted-by":"publisher","DOI":"10.1083\/jcb.17.2.375"},{"key":"e_1_3_1_2_4","first-page":"G467","volume":"32","author":"Anderson J. M.","year":"1995","unstructured":"Anderson J. M., Van Itallie C. M., Am. J. Physiol. 32, G467 (1995).","journal-title":"Am. J. Physiol."},{"key":"e_1_3_1_3_2","doi-asserted-by":"publisher","DOI":"10.1146\/annurev.physiol.60.1.143"},{"key":"e_1_3_1_4_2","unstructured":"L. L. Mitic and J. M. Anderson ibid. p. 121."},{"key":"e_1_3_1_5_2","doi-asserted-by":"publisher","DOI":"10.1172\/JCI110514"},{"key":"e_1_3_1_5_3","first-page":"157","volume":"16","author":"Di Stefano A.","year":"1993","unstructured":"Di Stefano A., Roinel N., De Rouffignac C., Wittner M., Renal Physiol. Biochem. 16, 157 (1993);","journal-title":"Renal Physiol. Biochem."},{"key":"e_1_3_1_5_4","doi-asserted-by":"publisher","DOI":"10.1152\/physrev.1994.74.2.305"},{"key":"e_1_3_1_6_2","doi-asserted-by":"publisher","DOI":"10.1152\/ajplegacy.1974.227.4.891"},{"key":"e_1_3_1_6_3","first-page":"58","volume":"18","author":"Kelepouris E.","year":"1998","unstructured":"Kelepouris E., Agus Z. S., Sem. Nephrol. 18, 58 (1998).","journal-title":"Sem. Nephrol."},{"key":"e_1_3_1_7_2","first-page":"F187","volume":"238","author":"Quamme G.","year":"1980","unstructured":"Quamme G., Dirks J. H., Am. J. Physiol. 238, F187 (1980).","journal-title":"Am. J. Physiol."},{"key":"e_1_3_1_8_2","first-page":"455","volume":"71","author":"Steele T. H.","year":"1968","unstructured":"Steele T. H., Wen S. F., Evenson M. A., Rieselbach R. E., J. Lab. Clin. Med. 71, 455 (1968).","journal-title":"J. Lab. Clin. Med."},{"key":"e_1_3_1_9_2","first-page":"25","volume":"33","author":"Kokko J. P.","year":"1974","unstructured":"Kokko J. P., Fed. Proc. 33, 25 (1974).","journal-title":"Fed. Proc."},{"key":"e_1_3_1_10_2","doi-asserted-by":"publisher","DOI":"10.1038\/ki.1995.199"},{"key":"e_1_3_1_10_3","unstructured":". This disease is number 248250 in the Online Mendelian Inheritance in Man (OMIM) database."},{"key":"e_1_3_1_11_2","unstructured":"Kindreds with recessive renal hypomagnesemia were recruited via ascertainment of affected index cases. Genomic DNA was prepared from venous blood of kindred members. The study was performed with approval of the Yale Human Investigation Committee."},{"key":"e_1_3_1_12_2","unstructured":"Additional data are available at www.sciencemag.org\/feature\/data\/1041171.shl."},{"key":"e_1_3_1_13_2","unstructured":"Genotyping was performed by polymerase chain reaction (PCR) using di- and tetranucleotide repeat polymorphisms. Analysis of linkage was performed by homozygosity mapping using the Genehunter program. The trait locus was specified as autosomal recessive with a mutant allele frequency of 0.002 99% penetrance and a phenocopy rate of 1%."},{"key":"e_1_3_1_14_2","unstructured":"BAC clones were selected from the RPCI 11 library by hybridization to STSs and their sizes determined by pulse-field gel electrophoresis. BAC ends were sequenced and overlaps of BAC clones were determined by STS content. Locus 539-5 a dinucleotide repeat with 85% heterozygosity was identified on BAC 4053-6 (11)."},{"key":"e_1_3_1_15_2","unstructured":"Exon trapping was performed by subcloning BACs into the vector pSPL3 (Gibco-BRL) transfection into COS-7 cells and isolation of spliced products which were cloned and sequenced. This identified exon 3 of PCLN-1 and the complete ORF (GenBank accession number ) was determined by selecting overlapping clones from a human kidney cDNA library. The deduced sequence was confirmed using specific primers to direct PCR from first-strand cDNA derived from human kidney."},{"key":"e_1_3_1_16_2","unstructured":"Analysis of hydrophobicity and sequence similarity was performed using Protean-Kyte-Doolittle hydropathy plot and Megalign Clustal method software respectively (DNAStar)."},{"key":"e_1_3_1_17_2","first-page":"5759","volume":"78","author":"Morita K.","year":"1999","unstructured":"Morita K., Furuse M., Fujimoto K., Tsukita S., Proc. Natl. Acad. Sci. U.S.A. 78, 5759 (1999).","journal-title":"Proc. Natl. Acad. Sci. U.S.A."},{"key":"e_1_3_1_18_2","doi-asserted-by":"publisher","DOI":"10.1083\/jcb.141.7.1539"},{"key":"e_1_3_1_19_2","doi-asserted-by":"publisher","DOI":"10.1126\/science.275.5296.73"},{"key":"e_1_3_1_20_2","unstructured":"Exons 3 to 5 of PCLN-1 cDNA were hybridized to human multiple-tissue Northern blots (Clontech) following standard protocols. A \u03b2-actin control probe was used separately to ensure RNA integrity (Y. Lu data not shown)."},{"key":"e_1_3_1_21_2","unstructured":"The intron-exon boundaries of PCLN-1 were determined by either amplification of gene segments by PCR or subcloning from BAC clones and sequencing of the products."},{"key":"e_1_3_1_22_2","unstructured":"Variants in PCLN-1 were identified using single-strand conformation polymorphism (SSCP) with PCLN-1 \u2013specific primer pairs directing PCR using genomic DNA as a template (11). Identified variants were sequenced."},{"key":"e_1_3_1_23_2","unstructured":"A segment of rabbit PCLN-1 cDNA (amino acids 99 to 192 in the human protein) was amplified from a rabbit kidney cDNA library (Clontech) using human PCLN-1 primers. The DNA sequence of the rabbit segment predicts a protein 90% identical to human PCLN-1. Reverse transcription PCR (RT-PCR) with microdissected rabbit nephron segments was performed as described ["},{"key":"e_1_3_1_23_3","doi-asserted-by":"publisher","DOI":"10.1046\/j.1523-1755.1998.00036.x"},{"key":"e_1_3_1_23_4","unstructured":"] using rabbit-specific PCLN-1 cDNA primers that lie in adjacent exons. Each segment was independently dissected five times and the results were identical each time with the exceptions that the DCT was negative once and the cortical collecting duct (CCD) was positive once."},{"key":"e_1_3_1_24_2","unstructured":"The peptide VSMAKSYSAPRTETAKMYAVD (29) from the COOH-terminus of PCLN-1 was coupled to keyhole limpet hemocyanin. Rabbits were immunized by successive injections and serum was harvested. Anti\u2013PCLN-1 was affinity-purified using the immunizing peptide coupled to an iodoacetyl cross-linked agarose column. Antibody specificity was determined by protein immunoblotting of bacterial lysates containing polyhistidine-tagged PCLN-1 in vector pET15b (Novagen). A product with an apparent molecular mass of 36 kD was detected in PCLN-1\u2013containing lysates (but not vector-only lysates) using anti\u2013PCLN-1 or mouse monoclonal anti-polyhistidine but not preimmune serum."},{"key":"e_1_3_1_25_2","doi-asserted-by":"publisher","DOI":"10.1083\/jcb.123.6.1777"},{"key":"e_1_3_1_25_3","unstructured":". Sections (5 \u03bcm) were cut from frozen human kidney specimens fixed and immunostained. Sections were visualized via immunofluorescence and confocal microscopy."},{"key":"e_1_3_1_26_2","doi-asserted-by":"publisher","DOI":"10.1038\/ng0196-24"},{"key":"e_1_3_1_27_2","doi-asserted-by":"publisher","DOI":"10.1093\/hmg\/6.9.1491"},{"key":"e_1_3_1_27_3","doi-asserted-by":"publisher","DOI":"10.1086\/302199"},{"key":"e_1_3_1_28_2","doi-asserted-by":"publisher","DOI":"10.1007\/BF01870332"},{"key":"e_1_3_1_29_2","doi-asserted-by":"publisher","DOI":"10.1146\/annurev.physiol.60.1.179"},{"key":"e_1_3_1_30_2","unstructured":"Abbreviations for the amino acid residues are as follows: A Ala; C Cys; D Asp; E Glu; F Phe; G Gly; H His; I Ile; K Lys; L Leu; M Met; N Asn; P Pro; Q Gln; R Arg; S Ser; T Thr; V Val; W Trp; and Y Tyr."},{"key":"e_1_3_1_31_2","unstructured":"K. A. Choate data not shown."},{"key":"e_1_3_1_32_2","unstructured":"We thank the families studied for their participation; V. Benigno S. Vittoria and M. Bianchetti for clinical data; C. Nelson-Williams for management of the DNA database; J. Anderson and P. De Camilli for thoughtful advice; and S. Frankel S. Floyd and W. Kim for help with confocal microscopy. Supported in part by NIH grant RO1DK51696. K.A.C. is an investigator of the NIH Medical Scientist Training Program; R.P.L. is an investigator of the Howard Hughes Medical Institute."}],"container-title":["Science"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.science.org\/doi\/pdf\/10.1126\/science.285.5424.103","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2024,1,13]],"date-time":"2024-01-13T09:02:32Z","timestamp":1705136552000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.science.org\/doi\/10.1126\/science.285.5424.103"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[1999,7,2]]},"references-count":41,"journal-issue":{"issue":"5424","published-print":{"date-parts":[[1999,7,2]]}},"alternative-id":["10.1126\/science.285.5424.103"],"URL":"https:\/\/doi.org\/10.1126\/science.285.5424.103","relation":{},"ISSN":["0036-8075","1095-9203"],"issn-type":[{"value":"0036-8075","type":"print"},{"value":"1095-9203","type":"electronic"}],"subject":[],"published":{"date-parts":[[1999,7,2]]}}}