Paracellin-1, a renal tight junction protein required for paracellular Mg2+ resorption

Science

Volumn 285, Issue 5424, 1999, Pages 103-106

  Simon, David B ^a   Lu, Yin ^a   Choate, Keith A ^a   Velazquez, Heino ^a   Al Sabban, Essam ^b   Praga, Manuel ^c   Casari, Giorgio ^d   Bettinelli, Alberto ^e   Colussi, Giacomo ^f   Rodriguez Soriano, Juan ^g   McCredie, David ^h   Milford, David ⁱ   Sanjad, Sami ^j   Lifton, Richard P ^a  

^a YALE UNIVERSITY SCHOOL OF MEDICINE   (United States)

^b KING FAISAL SPECIALIST HOSPITAL AND RESEARCH CENTER   (Saudi Arabia)

^c HOSPITAL UNIVERSITARIO 12 DE OCTUBRE   (Spain)

^d TELETHON INSTITUTE OF GENETICS AND MEDICINE TIGEM   (Italy)

^e UNIVERSITY OF MILAN   (Italy)

^f OSPEDALE NIGUARDA CA GRANDA   (Italy)

^g HOSPITAL DE CRUCES   (Spain)

^h ROYAL CHILDREN'S HOSPITAL   (Australia)

ⁱ BIRMINGHAM CHILDREN S HOSPITAL   (United Kingdom)

^j AMERICAN UNIVERSITY OF BEIRUT   (Lebanon)

more..

Author keywords

[No Author keywords available]

Indexed keywords

MAGNESIUM; PARACELLIN 1; PROTEIN; UNCLASSIFIED DRUG;

ARTICLE; CELL MEMBRANE; CONDUCTANCE; GENE EXPRESSION; GENE LOCUS; GENE MAPPING; HOMEOSTASIS; HYBRIDIZATION; HYPERCALCIURIA; HYPOMAGNESEMIA; MAMMAL; MEMBRANE PERMEABILITY; PRIORITY JOURNAL; PROTEIN FAMILY; RECESSIVE INHERITANCE; TIGHT JUNCTION;

AMINO ACID SEQUENCE; CALCIUM; CHROMOSOMES, HUMAN, PAIR 3; CLONING, MOLECULAR; FEMALE; GENES, RECESSIVE; HOMEOSTASIS; HUMANS; KIDNEY DISEASES; KIDNEY TUBULES; LOOP OF HENLE; MAGNESIUM; MAGNESIUM DEFICIENCY; MALE; MEMBRANE PROTEINS; MOLECULAR SEQUENCE DATA; MUTATION; PEDIGREE; PHYSICAL CHROMOSOME MAPPING; TIGHT JUNCTIONS;

MAMMALIA;

EID: 0033516683     PISSN: 00368075     EISSN: None     Source Type: Journal    
DOI: 10.1126/science.285.5424.103     Document Type: Article

References (37)

1. R. Bonnet, Dtsch. Med. Wochenschr. 21, 58 (1895); M. C. Farquhar and G. E. Palade, J. Cell Biol. 17, 375 (1963); J. M. Anderson and C. M. Van Itallie, Am. J. Physiol. 32, G467 (1995).
2. R. Bonnet, Dtsch. Med. Wochenschr. 21, 58 (1895); M. C. Farquhar and G. E. Palade, J. Cell Biol. 17, 375 (1963); J. M. Anderson and C. M. Van Itallie, Am. J. Physiol. 32, G467 (1995).
3. R. Bonnet, Dtsch. Med. Wochenschr. 21, 58 (1895); M. C. Farquhar and G. E. Palade, J. Cell Biol. 17, 375 (1963); J. M. Anderson and C. M. Van Itallie, Am. J. Physiol. 32, G467 (1995).
4. J. I. Madara, Annu. Rev. Physiol. 60, 143 (1998).
5. L. L Mitic and J. M. Anderson, ibid., p. 121.
6. G. R. Shareghi and Z. S. Agus, J. Clin. Invest. 69, 759 (1982); A. Di Stefano, N. Roinel, C. De Rouffignac, M. Wittner, Renal Physiol. Biochem. 16, 157 (1993); C De Rouffignac and G. Quamme, Physiol. Rev. 74, 305 (1994).
7. G. R. Shareghi and Z. S. Agus, J. Clin. Invest. 69, 759 (1982); A. Di Stefano, N. Roinel, C. De Rouffignac, M. Wittner, Renal Physiol. Biochem. 16, 157 (1993); C De Rouffignac and G. Quamme, Physiol. Rev. 74, 305 (1994).
8. G. R. Shareghi and Z. S. Agus, J. Clin. Invest. 69, 759 (1982); A. Di Stefano, N. Roinel, C. De Rouffignac, M. Wittner, Renal Physiol. Biochem. 16, 157 (1993); C De Rouffignac and G. Quamme, Physiol. Rev. 74, 305 (1994).
9. M. G. Brunette, N. Vigneault, S. Carriere, Am. J. Physiol. 227, 891 (1974); E. Kelepouris and Z. S. Agus, Sem. Nephrol. 18, 58 (1998).
10. M. G. Brunette, N. Vigneault, S. Carriere, Am. J. Physiol. 227, 891 (1974); E. Kelepouris and Z. S. Agus, Sem. Nephrol. 18, 58 (1998).
11. G. Quamme and J. H. Dirks, Am. J. Physiol. 238, F187 (1980).
12. T. H. Steele, S. F. Wen, M. A. Evenson, R. E. Rieselbach, J. Lab. Clin. Med. 71, 455 (1968).
13. J. P. Kokko, Fed. Proc. 33, 25 (1974).
14. M. Praga et al. Kidney Int. 47, 1419 (1995). This disease is number 248250 in the Online Mendelian Inheritance in Man (OMIM) database.
15. 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.
16. Additional data are available at www.sciencemag. org/feature/data/1041171.shl.
17. 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%.
18. 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).
19. 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 AF152101) 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.
20. Analysis of hydrophobicity and sequence similarity was performed using Protean-Kyte-Doolittle hydropathy plot and Megalign Clustal method software, respectively (DNAStar).
21. K. Morita, M. Furuse, K. Fujimoto, S. Tsukita, Proc. Natl. Acad. Sci. U.S.A. 78, 5759 (1999).
22. M. Furuse, K. Fujita, T. Hiragi, K. Fujimoto, S. Tsukita, J. Cell Biol. 141, 1539 (1998).
23. Z. Songyang et al., Science 275, 73 (1997).
24. Exons 3 to 5 of PCLW-1 cDNA were hybridized to human multiple-tissue Northern blots (Clontech) following standard protocols. A β-actin control probe was used separately to ensure RNA integrity (Y. Lu, data not shown).
25. 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.
26. Variants in PCLN-1 were identified using single-strand conformation polymorphism (SSCP) with PCLN-1-specific primer pairs directing PCR using genomic DNA as a template (11). Identified variants were sequenced.
27. 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 [H. Velazquez et al., Kidney Int. 54, 464 (1998)], using rabbit-specific PCLW-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.
28. 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-PCLN-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-containing lysates (but not vector-only lysates) using anti-PCLN-1 or mouse monoclonal anti-polyhistidine but not preimmune serum.
29. M. Furuse et al., J. Cell Biol. 123, 1777 (1993). Sections (5 μm) were cut from frozen human kidney specimens, fixed, and immunostained. Sections were visualized via immunofluorescence and confocal microscopy.
30. D. B. Simon et al., Nature Genet. 12, 24 (1996).
31. R. Y. Walder et al., Hum. Mol. Genet. 6, 1491 (1997); I. C. Meij et al., Am. J. Hum. Genet. 64, 180 (1999).
32. R. Y. Walder et al., Hum. Mol. Genet. 6, 1491 (1997); I. C. Meij et al., Am. J. Hum. Genet. 64, 180 (1999).
33. P. Claude, J. Membr. Biol. 39, 219 (1978).
34. P. A. Friedman, Annu. Rev. Physiol 60, 179 (1998).
35. 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.
36. K. A. Choate, data not shown.
37. 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.