Het ontraadselen van de moleculaire functie van androglobin: een testis specifiek globine
31 januari 2018
Promotiezaal (UAntwerpen, Campus Drie Eiken, Gebouw Q) - Universiteitsplein 1 - 2610 Wilrijk (Antwerpen) (route: UAntwerpen, Campus Drie Eiken
16 - 18 uur
Doctoraatsverdediging An Bracke - Departement Biomedische Wetenschappen
Androglobin (Adgb) is a recently discovered globin type consisting of large chimeric proteins with an N-terminal calpain-like protease domain and a central globin domain. Adgb is predominantly expressed in testis tissue and the expression is associated with the post-meiotic stages of spermatogenesis. Although we knew that Adgb plays a crucial role in male fertility, its molecular function is still not clear. The aim of this thesis was clear: unraveling the molecular function of Adgb. This was approached by using different strategies: the function was studied on biochemical, cellular and biological level.
To perform an in vitro biochemical characterization of the globin domain of Adgb, the globin domain was recombinantly expressed in E.coli, P.pastoris and baculovirus infected insect cells. We encountered difficulties with the solubility of the globin domain and concluded that for a proper folding it requires the context of the full length Adgb protein. Hence, we expressed the full length Adgb in insect cells and noticed that the protein was (auto-)cleaved over-time, which made the subsequent biochemical characterization of full length Adgb impossible. In future, these problems might be overcome by site-directed mutagenesis of the catalytic residue(s) of the calpain-like domain of Adgb (Chapter IV).
In order to unravel the cellular function of Adgb, we determined the protein-interaction partners of Adgb in testis tissue and we concluded that many of them are components of the chromatoid body, a specialized organelle in the male germ cell, of which it is assumed to serve as a center for mRNA storage and processing in the late, transcriptional-silent phases of spermatogenesis. Further (biochemical) characterization of these protein-protein interactions is required in order to unravel the molecular function of Adgb in this specialized organelle (Chapter V).
Finally, a biological analysis in human sperm samples and testis tissue exposed that Adgb expression is down-regulated or even absent in testis biopsies from men with disturbed or absent spermatogenesis. In future, more extended expression analysis in more testis biopsies, with different types of infertility pathologies is required to elucidate the function of Adgb in male infertility. Surprisingly, we discovered that it was impossible to detect Adgb on protein level in mature spermatozoa or in testis tissue. This may indicate that Adgb has a very transient role during spermatogenesis and that this low expression level will probably preclude the use of Adgb as a biomarker of male infertility (Chapter VI).