IBYME   02675
INSTITUTO DE BIOLOGIA Y MEDICINA EXPERIMENTAL
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
Presence of a GLIPR1 family member in the mouse epididymis
Autor/es:
ERNESTO JI; CALB D; VISCONTI PE; COHEN DJ; CUASNICU PS
Lugar:
San Pablo
Reunión:
Congreso; Fifth International Conference on Epididymis; 2010
Institución organizadora:
Maria Christina W. Avellar y Patricia S. Cuasnicu
Resumen:
The CAP superfamily includes four distinct groups: the cysteine-rich secretory proteins (CRISPs), the human glioma pathogenesis-related 1 (GLIPR1) proteins, the peptidase inhibitor 15 (Pi15) proteins, and the Golgi-associated pathogenesis-related 1 (GAPR-1) proteins. All members of the CAP superfamily are characterized by the presence of the   Pr-1 domain in the N amino terminal region, two signature sequence motifs within this domain, and a hinge region containing four cysteines.  The isolation and proteomic analysis of mouse sperm lipid rafts revealed the presence of two GLIPR1 family members, GLIPR1l1 and GLIPR1l2. Based on our reports on the role of CRISP proteins in different events of fertilization, in the present work we have cloned and characterized both Glipr1l1 and Glipr1l2 as a first step to study their potential role in the fertilization process. A sequence homology comparison revealed that all Glpr1 genes exhibit 30-80% identity and 50-90% homology. The study of the deduced protein sequences revealed that GLIPR1L1 is predicted to have a short C-terminal region lacking both a transmembrane domain and a glutamate rich domain (ERD) predicted to be present in the C-terminal region of GLIPR1L2. Studies of RT-PCR tissue screening showed high testicular expression for Glipr1l1 and expression in both the testis and caput epididymis for Glipr1l2. Semi-quantitative RT-PCR of Gliprl1 in the testis and of Glipr1l2 in both the testis and the epididymis showed a decrease in mRNA expression from day 0 (birth) to day 7, and a gradual increase thereafter reaching maximum levels at 60-70 days. Glpr1l1 and Glprll2 were cloned in E. coli, fused to His-tag. When the positive clones (selected by PCR screening) were subjected to IPTG induction, recombinant proteins with the predicted molecular weights of 30 kDa and 42 kDa for Gliprll1 and Gliprll2, respectively, were detected by SDS–PAGE analysis. In summary, we have characterized the tissue and developmental expression of Glipr1l1 and Glipr1l2 as well as successfully cloned and expressed the corresponding proteins. The availability of these recombinants proteins will allow the production of specific antibodies to investigate the potential role of GLIPR proteins in the maturation, capacitation and fertilization processes.