Unique and essential roles of RNA-binding proteins Musashi-1 and Musashi-2 during Spermatogenesis: Insights from a transgenic mouse model (#41)
Mammalian spermatogenesis is a complex developmental process requiring the strict regulation of stage-specific gene expression to produce physically mature spermatozoa from undifferentiated germ cells. Crucial to this process are RNA-binding proteins responsible for the regulated post-transcriptional control of essential mRNAs, particularly important during the two periods of inactive transcription which occur during spermatogenesis. Here we focus on the Musashi family of RNA binding proteins, in particular the role of mammalian Musashi-1 (MSI1) Musashi-2 (MSI2), both previously identified as key translational regulators in a variety of stem cell populations.
We determined, using a mouse model, that MSI1 is predominately expressed in gonocytes and later in spermatogonia, during the mitotic stages of division. In contrast MSI2 is not detected until the meiotic stages of germ cell development, in spermatocytes, and continues to be expressed through to round spermatid stage and during spermiogenesis.
Furthermore, we have extensively examined the role of MSI2 in spermatogenesis through the use of a transgenic mouse model, designed specifically to over-express full-length Msi2, in the developing testis. Completed breeding trials have demonstrated 100% sterility in MSI2 transgenic males, confirmed in vitro, and histological examination revealed aberrant spermatogenesis, significant depletion of post-meiotic germ cells, and major apoptosis/DNA damage.
Full-scale proteomic and microarray analyses undertaken on our transgenic MSI2 model has assisted in the characterisation of MSI2 interactions/targets. The microarray studies revealed the top biological functions corresponding to up-regulated molecules were cancer and cell death, with cell signalling and cell cycle among the top down-regulated functions, as confirmed via qPCR. Whilst differential proteomic analysis established that over-expression of Msi2 resulted in changes to a number of key transcriptional/translational regulators and several proteins crucial to mature sperm development and fertilisation.
These results provide conclusive evidence for the differential roles and unique functions of MSI1 and MSI2 RNA-binding proteins during germ cell development within the mammalian testis. Furthermore, based on the in-depth analysis of MSI2 through our novel transgenic model, we suggest that regulation of Msi2 is essential to normal spermatogenesis and crucial to fertility.