The sequence, expression, and chromosomal localization of a novel polycystic kidney disease 1-like gene, PKD1L1, in human

Polycystin-1 and polycystin-2 are the products of PKD1 and PKD2, genes that are mutated in most cases of autosomal dominant polycystic kidney disease. Since the first two polycystins were cloned, three new members, polycystin-L, -2L2, and -REJ, have been identified. In this study, we describe a sixth member of the family, polycystin-1L1, encoded by PKD1L1 in human. The full-length cDNA sequence of PKD1L1, determined from human testis cDNA, encodes a 2849-amino-acid protein and 58 exons in a 187-kb genomic region. The deduced amino acid sequence of polycystin-1L1 has significant homology with all known polycystins, but the longest stretches of homology were found with polycystin-1 and -REJ over the 1453- and 932-amino-acid residues, respectively. Polycystin-1L1 is predicted to have two Ig-like PKD, a REJ, a GPS, a LH2/PLAT, a coiled-coil, and 11 putative transmembrane domains. Several rhodopsin-like G-protein-coupled receptor (GPCR) signatures are also found in polycystin-1L1. Dot-blot analysis and RT-PCR revealed that human PKD1L1 is expressed in testis and in fetal and adult heart. In situ hybridization analysis showed that the most abundant and specific expression of Pkd1l1 was found in Leydig cells, a known source of testosterone production, in mouse testis. We have assigned PKD1L1 to the short arm of human chromosome 7 in bands p12--p13 and Pkd1l1 to mouse chromosome 11 in band A2 by fluorescence in situ hybridization. We hypothesize a role for polycystin-1L1 in the heart and in the male reproductive system.     

Molecular cloning of LMO41, a new human LIM domain gene

We have identified a new human LIM domain gene by isolating an autoantigenic cDNA clone from a human breast tumor cDNA library. The predicted amino acid sequence of the cDNA clone's 495 bp open reading frame contains two tandem LIM domain motifs, and within the LIM domain region there is 62% identity with the analogous region of the LIM-only gene LMO1. The homology to LMO1 is restricted to the 360 bp region encoding the tandemly repeated LIM domains, the rest of the open reading frame as well as the extensive, GC-rich 5' untranslated region, and 3' region of the 2 kb cDNA sequence are unrelated to any known genes. This gene has been designated LMO4.     

Molecular cloning, expression and chromosomal localization of a novel human REG family gene, REG III

Regenerating gene (Reg), first isolated from a regenerating islet cDNA library, encodes a secretory protein with a growth stimulating effect on pancreatic beta cells that ameliorates the diabetes of 90% depancreatized rats and non-obese diabetic mice. Reg and Reg-related genes have been revealed to constitute a multigene family, the Reg family, which consists of four subtypes (types I, II, III, IV) based on the primary structures of the encoded proteins of the genes [Diabetes 51(Suppl. 3) (2002) S462]. Plural type III Reg genes were found in mouse and rat. On the other hand, only one type III REG gene, HIP/PAP (gene expressed in hepatocellular carcinoma-intestine-pancreas/gene encoding pancreatitis-associated protein), was found in human. In the present study, we found a novel human type III REG gene, REG III. This gene is divided into six exons spanning about 3 kilobase pairs (kb), and encodes a 175 amino acid (aa) protein with 85% homology with HIP/PAP. REG III was expressed predominantly in pancreas and testis, but not in small intestine, whereas HIP/PAP was expressed strongly in pancreas and small intestine. IL-6 responsive elements existed in the 5'-upstream region of the human REG III gene indicating that the human REG III gene might be induced during acute pancreatitis. All the human REG family genes identified so far (REG Ialpha, REG Ibeta, HIP/PAP, REG III and REG IV) have a common gene structure with 6 exons and 5 introns, and encode homologous 158-175-aa secretory proteins. By database searching and PCR analysis using a yeast artificial chromosome clone, the human REG family genes on chromosome 2, except for REG IV on chromosome 1, were mapped to a contiguous 140 kb region of the human chromosome 2p12. The gene order from centromere to telomere was 5' HIP/PAP 3'-5' RS 3'-3' REG Ialpha 5'-5' REG Ibeta 3'-3' REG III 5'. These results suggest that the human REG gene family is constituted from an ancestor gene by gene duplication and forms a gene cluster on the region.     

Isolation and chromosomal localization of a novel FMS-like tyrosine kinase gene

We have isolated and sequenced part of a new gene of the tyrosine kinase family. This gene, called FLT3, has strong sequence similarities with members of a group of genes encoding growth factor receptors: FMS, KIT, and PDGFR. We have localized the human FLT3 gene to chromosome 13, band q12, and its mouse homolog to chromosome 5, region G.     

Nucleotide sequence, chromosomal localization and developmental expression of the mouse int-1-related gene

cDNA clones encoding the murine int-1-related protein (m-irp) were isolated from an 8.5-day mouse embryo library. m-irp and its human counterpart, h-irp, share extensive nucleotide homology in coding (92%) and 3' untranslated (69%) regions. At the amino acid level, m-irp and h-irp share 97% of amino acids including all 24 cysteine residues, which are highly conserved among members of the int-1 family. However, in contrast to h-irp and int-1, the predicted m-irp protein sequence did not contain a signal peptide sequence. Analysis of polymerase chain reaction, amplified cDNA, and genomic sequences strongly suggests that a single-base substitution has created a new 5' splice site 17 bp 5' of a highly conserved splice site. Splicing at this new site generates a mRNA-encoding an amino-terminal truncated protein. Splicing at the conserved splice site generates a mRNA species encoding a protein with a signal peptide sequence similar to h-irp. Close linkage between m-irp and the met oncogene maps m-irp sequences to proximal mouse chromosome 6. Adult and fetal expression of m-irp was examined by RNA blot analysis. Adult expression of m-irp is restricted to lungs and heart, and fetal expression, to placental tissue and to all stages of fetal development examined. In situ hybridization localized early fetal m-irp expression to the pericardium of the heart, to the umbilicus and associated allantoic mesoderm, and to the ventral lateral mesenchyme tissue surrounding the umbilical vein in the fetus. These results suggest a role for m-irp in the development of fetal allantoic communication.     

Structure and chromosomal localization of the mouse oncomodulin gene

The rat gene encoding oncomodulin (OM), a small calcium-binding protein, is under the control of a solo LTR derived from an endogenous intracisternal A-particle. The latter sequence is the only OM promoter analyzed so far. In order to study cell-type-specific OM expression in a species lacking LTR sequences in the OM locus, we initially synthesized an OM cDNA from mouse placenta. By sequencing, we found a 137-bp-long 5 leader region that differed markedly from its rat counterpart but had high similarity to several mouse genomic sequences. Primers specific to this sequence in addition with primers specific for an exon 2/intron 2 sequence were used to screen a mouse ES cell line genomic P1 library. One positive clone contained the whole OM gene, including intron 1 of 25 kb and a 5 flanking region of 27 kb lacking an LTR. The region upstream of exon 1 contains no TATA or CCAAT boxes but has a homopurine/homopyrimidine stretch of 102 bp as well as a (CA)₂₂ repeat. The latter sequence is polymorphic and was therefore, used to map the OM gene to the distal end of the long arm of mouse Chromosome (Chr) 5 by interspecific backcross analysis. Additonally we localized the OM gene by in situ hybridization to the region G1-3 on Chr 5, confirming the genetic linkage results. Finally, the OM gene was found to be structurally conserved and to exist in a single copy in mammals.     

Isolation and chromosomal localization of a novel nonerythroid ankyrin gene.

Immunoreactive isoforms of erythrocyte ankyrin have been shown to be present in a variety of nonerythroid tissues. Isolation of the genes that encode these isoforms will clarify their relationship to erythrocyte ankyrin. Using an erythrocyte ankyrin cDNA clone as a hybridization probe, we screened a human genomic library and isolated a clone that hybridizes with the probe at low stringency but not at high stringency. Partial nucleotide sequence of the clone revealed the presence of a 99-bp segment that is homologous to an exon of the erythrocyte ankyrin gene. Northern analysis showed that a labeled fragment of the clone hybridized to a 7-kb message in RNA of fetal brain but not of erythroid cells, suggesting that this clone is part of a novel gene that is expressed predominantly in nonerythroid tissue. Comparison of the sequence of the genomic clone with that of a recently isolated cDNA clone for brain ankyrin (Otto et al., 1989) showed identity of 96 of 99 bp between the putative exon and a segment of the cDNA clone (V. Bennett, personal communication, 1991), suggesting that the genomic clone is part of a gene for nonerythroid ankyrin, which we have designated ANK2. By analysis of somatic cell hybrids and fluorescence in situ hybridization, we assigned ANK2 to human chromosome 4 at a position equivalent to bands 4q25-q27.     

Identification of dynein heavy chain genes expressed in human and mouse testis: chromosomal localization of an axonemal dynein gene

Dynein heavy chains are involved in microtubule-dependent transport processes. While cytoplasmic dyneins are involved in chromosome or vesicle movement, axonemal dyneins are essential for motility of cilia and flagella. Here we report the isolation of dynein heavy chain (DHC)-like sequences in man and mouse. Using polymerase chain reaction and reverse-transcribed human and mouse testis RNA cDNA fragments encoding the conserved ATP binding region of dynein heavy chains were amplified. We identified 11 different mouse and eight human dynein-like sequences in testis which show high similarity to known dyneins of different species such as rat, sea urchin or green algae. Sequence similarities suggest that two of the mouse clones and one human clone encode putative cytoplasmic dynein heavy chains, whereas the other sequences show higher similarity to axonemal dyneins. Two of nine axonemal dynein isoforms identified in the mouse testis are more closely related to known outer arm dyneins, while seven clones seem to belong to the inner arm dynein group. Of the isolated human isoforms three clones were classified as outer arm and four clones as inner arm dynein heavy chains. Each of the DHC cDNAs corresponds to an individual gene as determined by Southern blot experiments. The alignment of the deduced protein sequences between human (HDHC) and mouse (MDHC) dynein fragments reveals higher similarity between single human and mouse sequences than between two sequences of the same species. Human and mouse cDNA fragments were used to isolate genomic clones. Two of these clones, gHDHC7 and gMDHC7, are homologous genes encoding axonemal inner arm dyneins. While the human clone is assigned to 3p21, the mouse gene maps to chromosome 14.     

cDNA sequence and chromosomal localization of the mouse parvalbumin gene, Pva

In the homozygous condition, the mutation adr (arrested development of righting response) of the mouse causes a myotonia and a drastic reduction of the Ca2+-binding protein parvalbumin (PV) in fast muscles. Using a rat PV probe, a mouse cDNA clone was isolated from a lambda gt11 wild-type fast-muscle library and its nucleotide sequence was determined. The protein coding and the 3' nontranslated regions of the mouse gene show extensive homology with the rat PV gene. The result of Southern blot hybridization is consistent with a single copy gene for parvalbumin. Restriction fragment length polymorphisms (RFLPs) between Mus musculus domesticus (e.g. C57BL/6) and Mus spretus (SPE) were detected with the enzymes Eco RI, Pst I, and Sst I. The restriction fragment patterns of DNA samples from 65 individual offspring of (C57BL/6 x SPE)F1 x C57BL/6 backcrosses were tested with the PV probe and matched, for linkage detection, to pre-existing patterns established with various RFLP probes on the same samples. A co-distribution of PV-RFLPs with Pvt-1 and Mlvi-2, which had been localized on chromosome 15, was detected. Thus, the structural gene for PV, designated Pva, maps to chromosome 15 of the mouse whereas the adr mutation shows no linkage with markers on this chromosome. Gene locus homology between chromosome 15 of the mouse and chromosome 22 of man (which carries the human PV gene) is discussed.