Protein Phosphatase 4 in human spermatozoa

: investigations into location, regulatory subunits and role in motility

Abstract

Infertility has been recognised as a public health issue worldwide by the WHO since 2004. The American Society of Reproductive Medicine cites 50% of infertility diagnosis as being of male factor origins. Surprisingly, there is very little that can be done to tackle the problem of male infertility. The key to progression in this field will be the translation of basic scientific research into the clinical laboratory. It is widely accepted that the process of maturation experienced by immature spermatozoa is the responsibility of post-translational modifications; a role of PKA in the regulation of sperm motility implies that protein phosphatases may be involved. The results presented in this thesis are the results of investigations into the role of serine/threonine protein phosphatases in human spermatozoa. It was hypothesised that the catalytic subunit of PPP4 would be present in human sperm and that it may play some role in the regulation of motility. Western Blotting was utilised to investigate whether the catalytic subunit of PPP4 was present in human spermatozoa; the 35kDa protein was present in all soluble fractions of spermatozoa prepared under both capacitating and non-capacitating conditions. Immunoprecipitation was performed to investigate what the catalytic subunit of PPP4 is associated with in human spermatozoa; AKAP4 was identified as a possible associate of PPP4c. Okadaic Acid was used to explore the effects on motility of the inhibition of PPP4 and PPP2A in human spermatozoa. The incubation of human spermatozoa with OA at concentrations aimed to inhibit PPP4 and PPP2A did not result in a significant increase or decrease to any of the motility parameters measured using the CASA. After incubation of human spermatozoa with OA at 100 fold the concentrations aimed to inhibit PPP4, significant increases in the percentage of progressively motile cells, ALH, BCF, STR and LIN were observed. The results presented here provide evidence for the first time of the presence of the catalytic subunit of PPP4 in human sperm and insights into its associated proteins. Evidence is also provided regarding the range of concentrations at which protein phosphatase inhibitors can be used to observe effects to the various motility parameters of human sperm.

Date of Award	2012
Original language	English
Awarding Institution	University of Dundee
Supervisor	Christopher Barratt (Supervisor)