The assembly of the sperm flagellum, the coordination of flagella formation with other events during spermiogenesis, the maintenance of flagellar structural integrity, and the regulation of the flagellar power stroke and waveform are poorly understood at the molecular level. The long-term goal of this research is to characterize regulatory proteins of the mammalian sperm axoneme, and to identify their roles in governing flagellar structure and activity. This knowledge will provide a molecular framework for understanding sperm motility defects causing male infertility and the function of """"""""9+2"""""""" cilia in general.
Four specific aims are proposed based upon our observations suggesting bi-functional roles for the Pf20 gene. Pf20 encodes two proteins, the 71 kDa PF20I, localized to the axoneme central apparatus, and a second protein transcribed from a testis-specific promoter, 35 kDa PF20s, which contains the C-terminus of PF20I, but is localized to the germ cell nucleus. PF20I is produced meiotically, PF20s accumulates post-meiotically. Highly chimeric mice with a mutated Pf20 allele that affects PF20s expression have a phenotype of severe oligospermia and sperm axonemal defects. These observations suggest that PF20I and PF20s may have distinct roles. PF20I in the axoneme interacts with SPAG6, apparently facilitating assembly or imparting stability to the sperm flagellar axoneme. PF20s interacts with a testis-specific serine threonine kinase, TSSK-2, and a chromosome-binding protein in the nucleus, meiosis induced gene 1 (MEIG1). Experiments will be conducted to establish that Pf20 is a unique gene that is involved in the formation/stability of an organelle (the axoneme) essential for sperm function (motility) and differentiation events (gene expression). The hypotheses to be tested include: 1) that PF20I plays an important role in sperm motility such that male mice lacking PF20I will be infertile due to a severe sperm motility defect due to abnormalities in the assembly or structural integrity of the sperm axoneme;2) that PF20s is required for normal post-meiotic sperm development, acting in the nucleus to regulate gene expression such that mice deficient in PF20s will be sterile due to post-meiotic spermatogenic failure;3) that TSSK-mediated phosphorylation regulates PF20s function;and 4) that PF20 mutations cause abnormalities in human spermatogenesis as well as primary ciliary dyskinesia. The latter hypothesis is supported by the discovery of oligospermic males heterozygous for mutations that would affect PF20s expression.
Showing the most recent 10 out of 24 publications
