RESUMEN
A novel human cDNA sequence has been isolated from human testis cDNA library. This sequence, named PD1, reveals an open reading frame encoding a protein of 520 amino acids. A partial sequence similarity has been found with the RBM gene involved in the regulation of human spermatogenesis. Northern blot analysis for PD1 mRNA from several human tissues demonstrated two distinct transcripts of 2.7 (more abundant) and 4.0 kb and revealed that PD1 is expressed in testis and to a lesser extent also in spleen, thymus, and prostate. Immunohistochemical analysis of human testis showed that this protein is detected in the cytoplasm of Sertoli cells. Antibodies against a rhPD1 fragment were used for Western blot analysis, which confirmed the presence of a 60-kDa molecule in crude extract of human testicular cells obtained from fine-needle aspiration and showed different patterns in various testiculopathies, suggesting a role for such gene in human spermatogenesis.
Asunto(s)
Proteínas/genética , Testículo/química , Testículo/patología , Secuencia de Aminoácidos , Secuencia de Bases , Northern Blotting , Southern Blotting , Clonación Molecular , ADN Complementario/genética , Humanos , Masculino , Datos de Secuencia Molecular , Proteínas Nucleares , Proteínas/aislamiento & purificación , Proteínas de Unión al ARN/genética , Túbulos Seminíferos/química , Homología de Secuencia de Aminoácido , Células de Sertoli/química , Recuento de Espermatozoides , Espermatogénesis , Distribución TisularRESUMEN
A novel 44-kDa gene product (D123) has been proposed as necessary for S-phase entry of the cell cycle: a point mutation resulted in a temperature-sensitive arrest in G1-phase. From human fibrosarcoma cDNA library, we have isolated an identical gene and studied its sequence and mRNA and protein expression. Compared with D123, three nucleotide differences within the human coding sequence, plus others, result in a change of two amino acids. A partial sequence similarity has been found with a yeast gene of unknown function. The protein has several potential phosphorylation sites, is highly hydrophilic, and may be highly structured in alpha-helix. The mRNA is abundantly expressed by a variety of normal and transformed cells and by all tissues examined, being most highly expressed in testis. Specific antibodies, raised against a rhD123 polypeptide, recognize a major 42- to 44-kDa molecule in crude extract of various human cell lines. Immunohistochemistry reveals that D123 protein is not homogeneously expressed: it is detected, often in granular vescicles, in the cytoplasm of some epithelial, stromal, and sperm cells and in varicosities lining nervous fibers, while it appears to be absent in nuclei, endothelial, and smooth muscle cells. The precise link between cytoplasmic occurrence of D123 and cell cycle progression still remains to be clarified.
