Liquid semen storage-induced alteration in the protein composition of turkey (Meleagris gallopavo) spermatozoa.

Liquid storage of turkey semen without the loss of fertilizing ability is of practical interest to the poultry industry. However, fertility rates from liquid-stored turkey semen decline within a few hours. A clear cause of the decline in spermatozoa quality remains unidentified. Therefore, the purpose of the present study was to monitor the dynamics of proteomic changes in spermatozoa during 48 h of liquid storage by 2-dimensional difference in-gel electrophoresis coupled with matrix-assisted laser desorption/ionization mass spectrometry. A total of 57 protein spots were differentially expressed between fresh and stored spermatozoa; 42 spots were more and 15 were less abundant after 48 h of semen storage. Raw proteomic data are available via ProteomeXchange with identifier PXD043050. The selected differentially expressed proteins (DEPs) were validated by western blotting and localized in specific spermatozoa structures by immunofluorescence, such as the head (acrosin and tubulin α), midpiece (acrosin, aconitate hydratase 2, and glycerol-3-phosphate dehydrogenase) and tail (tubulin α). Most of the DEPs that changed in response to liquid storage were related to flagellum-dependent cell motility, energy derivation through oxidation of organic compounds and induction of fertilization, suggesting the complexity of the processes leading to the decrease in stored semen quality. The damaging effect of liquid storage on spermatozoa flagellum manifested as more microtubule proteins, such as tubulins and tektins, most likely formed by posttranslational modifications, tubulin α relocation from the tail to the sperm head, which appeared after 48 h of semen storage, and decreases in fibrous shelf proteins at the same time. Motility could be affected by dysregulation of Ca2+-binding proteins and disturbances in energy metabolism in spermatozoa flagellum. Regarding sperm mitochondria, DEPs involved in energy derivation through the oxidation of organic compounds indicated disturbances in fatty acid beta oxidation and the tricarboxylic acid cycle as possible reasons for energy deficiency during liquid storage. Disturbances in acrosin and 1-phosphatidylinositol 4,5-bisphosphate phosphodiesterase zeta may be involved in rapid declines in the fertility potential of stored turkey spermatozoa. These results showed the complexity of the processes leading to a decrease in stored semen quality and broadened knowledge of the detrimental effects of liquid storage on turkey spermatozoa physiology.

The acquisition of desiccation tolerance in developing Vicia hirsuta seeds coincides with an increase in galactinol synthase expression and soluble α-D-galactosides accumulation.

Galactinol is the galactosyl donor for the biosynthesis of both the raffinose family oligosaccharides (RFOs) and galactosyl cyclitols (Gal-C). Its synthesis by galactinol synthase (GolS, EC 2.4.1.123) is the first committed step of the soluble α-D-galactosides biosynthetic pathway in orthodox seeds. The deposition of galactosides in seeds is suggested to be associated with desiccation tolerance (DT). In this work, for the first time, we cloned and characterized two Vicia hirsuta (L.) S.F. Gray galactinol synthase genes (VhGolS1, VhGolS2), analyzed galactinol synthase activity and measured the accumulation of galactosides of both sucrose and D-pinitol in relation to the acquisition of DT in developing seeds of this wild species. A developmentally induced increase of VhGolS1 expression preceded the rise of GolS activity in crude protein extract from maturing seeds, while the expression of the VhGolS2 gene remained low. GolS activity peaked just after the beginning of the maturation drying phase. The increase of GolS activity was not followed by galactinol accumulation, instead the high enzyme activity was related to high levels of galactose bound in soluble galactosides of the RFO and galactosyl pinitol series. Acquisition of DT coincided with an increase of VhGolS1 expression, high galactinol synthase activity and the accumulation of oligogalactosides in seeds. DT was positively correlated with the high content of soluble α-D-galactosides of both the RFO and galactosyl pinitol series as well as with the amount of galactose bound in these galactosides.