CENTRIN DYNAMICS IN THE MALE REPRODUCTIVE CELL

Project: Research project

Description

The sperm centrosome is intimately associated with organization and
morphogenesis of the spermatocyte, particularly with regard to
establishment of extreme cell polarity in mature sperm. In addition, the
sperm centrosome is delivered to the oocyte at the time of fertilization
where it plays a fundamental role in the organization of the cytoskeleton
of the zygote and early embryo through the formation of the
microtubule-based sperm aster and the first mitotic spindle. We have
recently established that centrin, a 21 kDa calcium-binding protein found
in centrosome-associated fibers, is a prominent component of the sperm
centrosome. Previous studies in other systems demonstrate a role for
centrin-based fibers in centrosome/mitotic spindle pole positioning,
segregation, and reorientation. Based on these observations, we present
the specific hypothesis that sperm centrin is an essential component of the
sperm connecting piece which functions in centrosome dynamics during sperm
morphogenesis, and in zygotes and early embryos during spindle assembly and
function. The work outlined in this proposal is directed toward achieving
a greater understanding of centrosome dynamics in the male reproductive
cell during spermatocyte differentiation and following fertilization of the
oocyte. For a thorough understanding of the contribution by sperm to the
zygotic centrosome it is important to determine the components of the sperm
connecting piece that are essential to the process of spermatocyte
differentiation, and sperm aster and centrosome formation and function.
What proteins are carried by the sperm connecting piece to the egg? Are
they essential for centrosome function? What is their relationship to
preexisting pools of centrosomal components present in the oocyte? Can
their function be perturbed by specific antibodies and/or drug treatments?
Our laboratory has developed immunological, biochemical, and molecular
tools which will allow us to readily probe the role of a recently
discovered and novel calcium-binding cytoskeletal protein, centrin, in
these processes. The specific aims of the proposed studies include; 1)
characterization of molecular and biochemical properties of sperm centrin
through the purification, peptide and epitope mapping, partial peptide
sequencing of centrin from sperm, and the molecular cloning of sperm
(testicular) centrin. 2) To define the precise structural organization of
centrin in developing spermatids, mature sperm, and in fertilized eggs and
early embryos by immunofluorescence and immunoelectron microscopy using
polyclonal and monoclonal antibodies raised against centrin. And 3), to
determine if i centrin plays an essential role in sperm aster formation and
centrosome behavior in early fertilized eggs. Finally, the proposed
studies are likely to shed light on the nature of several human infertility
disease states that involve centrosomal and spermatid malformations
including 'decapitated' sperm.
StatusFinished
Effective start/end date8/1/927/31/96

Funding

  • National Institutes of Health
  • National Institutes of Health
  • National Institutes of Health
  • National Institutes of Health

Fingerprint

centrosomes
germ cells
spermatozoa
calcium-binding proteins
spermatocytes
embryo (animal)
mitotic spindle apparatus
spermatids
zygote
oocytes
peptide mapping
cytoskeletal proteins
fluorescent antibody technique
drug therapy
molecular cloning
microscopy
monoclonal antibodies
antibodies

ASJC

  • Medicine(all)