The process of screening cDNA expression libraries with phageoptimized peptide ligands, termed cloning of ligand targets (COLT), was recently used to isolate a series of SH3 domaincontaining proteins (Sparks et al. 1996). Among the 18 SH3 domain-containing proteins identified, nine were previously unreported from mouse. The structures of seven of the proteins whose genes are mapped in this study are diagrammed in Fig. 1. The function of many of the mouse proteins is being actively pursued by many laboratories. Three of the proteins, SH3P4, SH3P8, and SH3P14, are highly related in structure and have been discovered to bind synaptojanin and dynamin, and to be involved in endocytosis (de Heuvel et al. 1997; Ringstad et al. 1997). This family of proteins, which has also been discovered in human (Giachino et al. 1997), has been termed endophilin 1, 2, and 3. SH3P8 was also identified as a gene fused to MLL in human acute myeloid leukemia (So et al. 1997) and as a protein that binds to the Gag protein of murine leukemia viruses in a yeast two-hybrid screen (W. Kim, T. Torrey, H. Morse, unpublished observations). SH3P9, renamed amphiphysin II because of its strong similarity to amphiphysin, has been shown to be a component of the endocytic machinery (Butler et al. 1997; Ramjaun et al. 1997) as well as the cytoskeleton, where it may regulate the c-Abl tyrosine kinase (Kadlec and Pendergast 1997). SH3P12 has been observed to interact with c-Cbl in cells where it may play a role in both signal transduction pathways and regulation of the cytoskeleton (Ribon et al. 1998). To learn more about the genes for the seven novel mouse proteins, we set out to map their chromosomal locations. Two sets of multilocus genetic crosses were analyzed for inheritance of the mouse genes encoding the SH3 family genes: (NFS/N or C58/J × M. m. musculus) × M. m. musculus (NMM and CMM; Kozak et al. 1990) and (NFS/N x M. spretus) × M. spretus or C58/J (NSS and NSC; Adamson et al. 1991). Probes were prepared by polymerase chain reaction (PCR) from the nine cDNA clones, seven of which have been deemed full-length (Sparks et al. 1996), and used in blots of restriction enzyme digests of genomic DNA. Inheritance patterns were described for eight polymorphic fragments identified by the seven different probes. Restriction fragments used to identify specific loci are given in Table 1. Seven loci were typed in the M. spretus crosses and six loci in the M. m. musculus crosses. Comparisons of the inheritance patterns of these fragments with those of over 1200 loci previously typed and mapped in these crosses showed that eight loci could be positioned on seven chromosomes. Two genes were mapped to Chromosome (Chr) 19, but they were separated by more than 20 cM. The recombination data used to derive the map locations are given in Table 2. In all but one case, all restriction fragments identified by a single probe could be mapped to the same locus. The exception, probe SH3P13, identified loci on two mouse chromosomes, Chrs 2 and 7. The two corresponding loci, Sh3d2c1 and Sh3d2c2, were identified in both sets of crosses, indicating that neither represented a species-specific pseudogene. Previous data have shown that the human homolog of this SH3 domain gene, originally termed SH3GL3, is located at 15q24 (Giachino et al. 1997). One of the mouse loci identified by this probe, Sh3d2c2, mapped to a region of Chr 7 homologous to 15q21-26, identifying it as the mouse homolog of human SH3GL3. The nature of the second locus, Sh3d2c, remains to be determined. Human map locations have also been defined for Sh3d2a and Sh3d2b. The homolog of Sh3d2a (SH3GL2) was mapped to 9p22, which is consistent with its position on mouse Chr 4. We previously reported the map location for Sh3d2b on mouse Chr 17 (Torrey et al. 1998), consistent with the human location for SH3GL1 on 19p13.3. Finally, another SH3 domain gene, Amph (amphiphysin), maps to mouse Chr 13 and human 7p14-p13 (Jenkins et al. 1995). None of the other genes have been mapped in human, although mouse map locations predict positions for SH3D4 and SH3D5 on human chromosomes 8p21-23 and 10q respectively. Since map locations for Dbn1l and Sh3d3 place them in regions with homology to two or more human chromosomes, we attempted to make