Proviral inactivation of the Npat gene of Mpv 20 mice results in early embryonic arrest.

The Mpv 20 transgenic mouse strain was created by infection of embryos with a defective retrovirus. When Mpv 20 heterozygous animals were crossed, no homozygous neonatal mice or midgestation embryos were identified. When embryos from heterozygous crosses were cultured in vitro, approximately one quarter arrested as uncompacted eight-cell embryos, indicating that proviral insertion resulted in a recessive lethal defect whose phenotype was manifest very early in development. Molecular cloning of the Mpv 20 insertion site revealed that the provirus had disrupted the Npat gene, a gene of unknown function, resulting in the production of a truncated Npat mRNA. Expression of the closely linked Atm gene was found to be unaffected by the provirus.

Genomic structure of Unp, a murine gene encoding a ubiquitin-specific protease.

The murine Unp gene encodes a ubiquitin-specific protease, a member of a family of enzymes that includes the product of the human tre-2 oncogene. The Unp gene has previously been mapped to chromosome 9. We have cloned in bacteriophage a 50 kilobase region of chromosome 9 containing the Unp gene, and have determined the nucleotide sequence of the gene. The gene has 22 exons, distributed over 47.4 kb. A processed ribosomal S2 pseudogene was identified in the third intron of the Unp gene. Expression of Unp is driven by a GC-rich, 'housekeeping' type promoter.

Molecular cloning of three cDNAs that encode cysteine proteinases in the digestive gland of the American lobster (Homarus americanus).

Three clones were isolated from a lobster digestive gland cDNA library, using oligonucleotide probes based on the partial amino terminal sequence of a digestive cysteine proteinase. The cDNAs, LCP1, LCP2 and LCP3 encode preproenzymes of 322, 323 and 321 amino acid residues, and putative mature enzymes of 217, 216 and 215 residues, respectively. Calculated mature protein molecular masses are 23386 (LCP1), 29093 (LCP2) and 23255 (LCP3) Sequence alignments show that the lobster enzymes are more similar to L (55-62% identity) than H (42-44%) or B (22-24%) cathepsins. Southern analysis indicated as many as eleven genes related to the three cDNAs.

Characterization of Sam68-like mammalian proteins SLM-1 and SLM-2: SLM-1 is a Src substrate during mitosis.

Sam68, the 68-kDa Src substrate associated during mitosis, is an RNA-binding protein with signaling properties that contains a GSG (GRP33, Sam68, GLD-1) domain. Here we report the cloning of two Sam68-like-mammalian proteins, SLM-1 and SLM-2. These proteins have an approximately 70% sequence identity with Sam68 in their GSG domain. SLM-1 and SLM-2 have the characteristic Sam68 SH2 and SH3 domain binding sites. SLM-1 is an RNA-binding protein that is tyrosine phosphorylated by Src during mitosis. SLM-1 bound the SH2 and SH3 domains of p59(fyn), Grb-2, phospholipase Cgamma-1 (PLCgamma-1), and/or p120(rasGAP), suggesting it may function as a multifunctional adapter protein for Src during mitosis. SLM-2 is an RNA-binding protein that is not tyrosine phosphorylated by Src or p59(fyn). Moreover, SLM-2 did not associate with the SH3 domains of p59(fyn), Grb-2, PLCgamma-1, or p120(rasGAP), suggesting that SLM-2 may not function as an adapter protein for these proteins. The identification of SLM-1 and SLM-2 demonstrates the presence of a Sam68/SLM family whose members have the potential to link signaling pathways with RNA metabolism.

The identification of two Drosophila K homology domain proteins. Kep1 and SAM are members of the Sam68 family of GSG domain proteins.

Sam68 is a member of a growing family of RNA-binding proteins that contains an extended K homology (KH) domain embedded in a larger domain called the GSG (GRP33, Sam68, GLD1) domain. To identify GSG domain family members, we searched data bases for expressed sequence tags encoding related portions of the Sam68 KH domain. Here we report the identification of two novel Drosophila KH domain proteins, which we termed KEP1 (KH encompassing protein) and SAM. SAM bears sequence identity with mammalian Sam68 and may be the Drosophila Sam68 homolog. We demonstrate that SAM, KEP1, and the recently identified Drosophila Who/How are RNA-binding proteins that are able to self-associate into homomultimers. The GSG domain of KEP1 and SAM was necessary to mediate the RNA binding and self-association. To elucidate the cellular roles of these proteins, SAM, KEP1, and Who/How were expressed in mammalian and Drosophila S2 cells. KEP1 and Who/How were nuclear and SAM was cytoplasmic. The expression of KEP1 and SAM, but not Who/How, activated apoptotic pathways in Drosophila S2 cells. The identification of KEP1 and SAM implies that a large GSG domain protein family exists and helps redefine the boundaries of the GSG domain. Taken together, our data suggest that KEP1 and SAM may play a role in the activation or regulation of apoptosis and further implicate the GSG domain in RNA binding and oligomerization.