Scott M. Silbiger

Neu differentiation factor: A transmembrane glycoprotein containing an EGF domain and an immunoglobulin homology unit

We recently reported that a 44 kd glycoprotein secreted by transformed fibroblasts stimulates tyrosine phosphorylation of the product of the neu proto-oncogene and induces differentiation of mammary tumor cells to milk-producing, growth-arrested cells. A partial amino acid sequence of the protein, termed Neu differentiation factor (NDF), enabled cloning of the corresponding complementary DNA. The deduced structure of the precursor of NDF indicated that it is a transmembrane protein whose extracellular portion contains an EGF-like domain that probably functions as a receptor recognition site. In addition, the ectodomain contains one immunoglobulin homology unit. Despite the lack of a recognizable hydrophobic signal peptide at the N-terminus, a recombinant NDF, like the natural molecule, is released into the medium of transfected COS-7 cells in a biologically active form. Northern blot analysis indicated the existence of several NDF transcripts, the major ones being 1.8, 2.6, and 6.7 kb in size. Transformation by the ras oncogene dramatically elevated the expression of NDF in fibroblasts.

Cloning of cDNAs encoding human TIMP-3, a novel member of the tissue inhibitor of metalloproteinase family.

Proteins of the tissue inhibitor of metalloproteinase (TIMP) family bind and inactivate matrix metalloproteinases such as collagenases and gelatinases. We report the cloning and sequencing of cDNAs encoding a novel human TIMP, which we designated TIMP-3, the third member of the human TIMP family. Degenerate PCR primers derived from highly conserved regions of TIMP family cDNAs amplified a 402-bp product from human fetal kidney cDNA. This product and a related 333-bp PCR product were used as probes to screen two cDNA libraries. Three TIMP-3 cDNA clones were isolated, including a 1240-bp fetal kidney clone that contained a complete TIMP-3 precursor coding region of 211 amino acids (aa). The deduced precursor protein includes twelve Cys and 27 other aa that are invariant in the TIMP family. The predicted aa sequence is 89, 39 and 46% identical to those of ChIMP-3, human TIMP-1 and human TIMP-2, respectively. Northern blot analyses detected three TIMP-3 mRNA bands of 2.2, 2.5 and 4.4 kb in several human cell lines.

Protein tyrosine phosphatase (PC12, Br7,Sl) family: Expression characterization in the adult human and mouse

Protein tyrosine phosphatases (PTPs) play important roles in modulating signals transduced by tyrosine kinases. Certain phosphatases have been implicated as having important roles in embryonic development as well as in adult physiology. Although both kinases and phosphatases are equally important in regulating signal transduction, phosphatases as a group have not been well characterized. Thus, characterization of sequence, expression, and biological function for additional phosphatases is informative. PTPBr7/PC12 and PTPSl are mouse receptor PTPs sharing similar amino acid sequences. Northern blot analysis demonstrated expression of these genes in adult rodent brain and revealed previously uncharacterized transcripts in the brain and other tissues. Our results demonstrate that PTPBr7/PC12 and PTPSl are members of a larger family of PTPs. We have identified two novel family members as well as several novel transcriptional splice variants from both human and mouse colon cDNA libraries. Expression analysis demonstrated that the various mRNA transcripts are differentially expressed, with the highest levels found in the brain, intestinal tract, uterus, and placenta. In situ hybridization analysis of mouse brain and intestinal tissues established that each isoform has a unique expression pattern in specific cell populations as well as in tissue regions. Furthermore, these restricted patterns suggest that the encoded family of phosphatases may play roles in modulating signal transduction pathways important for specific cell types and biological processes. Anat Rec 258:221–234, 2000.

Cloning and partial structure of the gene encoding human tissue inhibitor of metalloproteinases-3.

Using a cDNA probe, two genomic clones were obtained encoding the human tissue inhibitor of metalloproteinases-3 (TIMP-3). Analysis of these clones showed that they contained four distal exons and three introns of the gene. Although the intron-exon structure is similar to that of the timp1 gene, the first intron of the timp3 gene is much longer, being at least 17.5 kb in size.