Publicação
Somatic expression of the testis-specific PDHA2 gene : mechanisms of activation/silencing
| Resumo: | During spermatogenesis, germ cells undergo a complex process of cell differentiation and morphological restructuring, which depends on the coordinated expression of different genes. Some vital examples are those involved in cell energy metabolism, namely the genes encoding the E1α subunit of pyruvate dehydrogenase complex: the somatic PDHA1 (X-linked) and the testis-specific PDHA2 (autosomal). There are no data related to the study at the RNA and protein levels of PDHA genes during spermatogenesis. The present study aimed to describe the mRNA and protein expression patterns of the human PDHA genes during spermatogenesis. Expression profiles of the PDHA1 and PDHA2 genes were characterized using different human tissues and cells. Diploid and haploid germ cells fractions were obtained from testis tissues. The mRNA profiles were analyzed by qRT-PCR methods, whereas the protein profiles were evaluated by immunohistochemistry, western blotting and two-dimensional electrophoresis. Both PDHA proteins appeared localized to the cytoplasm. All somatic tissues showed similar levels of PDHA protein. Testis tissue, comprising somatic and germ cells, presented an increased protein level. PDHA2 was the sole protein species present in ejaculated spermatozoa. The developmental stage at which occurs the switch from PDHA1 to PDHA2 expression was determined by the study of mRNA expression levels during spermatogenesis, namely in diploid and haploid germ cells fractions, as well as in ejaculated spermatozoa. Expression of the PDHA1 gene was found in all somatic cells, whereas expression of PDHA2 gene was restricted to germ cells. The switch from the X-linked to the autosomic gene expression occurred in spermatocytes. Data suggests the activation of PDHA2 gene expression is most probably a mechanism to ensure the continued expression of the protein, thus allowing germ cell viability and functionality. |
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| Autores principais: | Pinheiro, Ana Sofia da Costa, 1981- |
| Assunto: | Teses de doutoramento - 2012 |
| Ano: | 2012 |
| País: | Portugal |
| Tipo de documento: | tese de doutoramento |
| Tipo de acesso: | acesso aberto |
| Instituição associada: | Universidade de Lisboa |
| Idioma: | inglês |
| Origem: | Repositório da Universidade de Lisboa |
| Resumo: | During spermatogenesis, germ cells undergo a complex process of cell differentiation and morphological restructuring, which depends on the coordinated expression of different genes. Some vital examples are those involved in cell energy metabolism, namely the genes encoding the E1α subunit of pyruvate dehydrogenase complex: the somatic PDHA1 (X-linked) and the testis-specific PDHA2 (autosomal). There are no data related to the study at the RNA and protein levels of PDHA genes during spermatogenesis. The present study aimed to describe the mRNA and protein expression patterns of the human PDHA genes during spermatogenesis. Expression profiles of the PDHA1 and PDHA2 genes were characterized using different human tissues and cells. Diploid and haploid germ cells fractions were obtained from testis tissues. The mRNA profiles were analyzed by qRT-PCR methods, whereas the protein profiles were evaluated by immunohistochemistry, western blotting and two-dimensional electrophoresis. Both PDHA proteins appeared localized to the cytoplasm. All somatic tissues showed similar levels of PDHA protein. Testis tissue, comprising somatic and germ cells, presented an increased protein level. PDHA2 was the sole protein species present in ejaculated spermatozoa. The developmental stage at which occurs the switch from PDHA1 to PDHA2 expression was determined by the study of mRNA expression levels during spermatogenesis, namely in diploid and haploid germ cells fractions, as well as in ejaculated spermatozoa. Expression of the PDHA1 gene was found in all somatic cells, whereas expression of PDHA2 gene was restricted to germ cells. The switch from the X-linked to the autosomic gene expression occurred in spermatocytes. Data suggests the activation of PDHA2 gene expression is most probably a mechanism to ensure the continued expression of the protein, thus allowing germ cell viability and functionality. |
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