Abstract/Details

Characterization of the sperm factor responsible for initiating [calcium (2+)](i) oscillations during fertilization in mammalian eggs


2000 2000

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Abstract (summary)

During fertilization the sperm activates the mammalian egg by eliciting [Ca2+]i oscillations. However, how the sperm triggers [Ca2+]i oscillations in mammalian eggs remains unknown. To test the possibility that a factors) from the sperm is able to elicit [Ca 2+]i oscillations, sperm fractions (factors) were prepared from different species and injected into mammalian oocytes or eggs. The results show that injection of sperm factor from either porcine or human sperm triggered long-lasting [Ca2+]i oscillations in mouse oocytes and bovine eggs in a pattern similar to the physiological responses observed in each of these species, implying that the sperm factor is functionally conserved among mammalian species. In addition, sperm factor-induced [Ca2+] i oscillations appeared to be mediated by the IP3 receptor, while the ryanodine receptor may be involved in the modulation of these oscillations. Furthermore, [Ca2+]i oscillations induced by sperm factor are capable of initiating normal egg activation and parthenogenetic development.

In order to isolate and characterize the unknown active molecule(s) in sperm factor, different tissues or cell extracts were screened for the presence of sperm factor like activity. It appears that the [Ca2+] i oscillation-inducing activity is sperm/testis specific. In addition, the results showed that the active component contains a protein moeity and that the same single active component of porcine sperm factor is present in both soluble and in less soluble sperm compartments. We also demonstrated that gpd/oscillin, a proposed active component of mammalian sperm factor, is not responsible for the [Ca2+]i oscillation-inducing activity as shown by the lack of effects on Ca2+ responses or absence in active fractions following immunodepletion or a combination of fractionation techniques. Similarly, our results also showed that neither PLCγ1, PLCγ2 nor tr-c-Kit is likely to be the active component of sperm factor.

Although we were unable to identify a specific candidate molecule, these experiments have led to the identification of three polypeptides in the final active fraction after sequential chromatographic steps. Our results also show that the active component has an isoelectric point of 6.5–7.0 and a relative molecular weight ranging from 29–68 kDa. These polypeptides will be sequenced and used to raise monoclonal or polyclonal antibodies, which will be used to screen a testis cDNA library. This last step is expected to lead to the isolation of the gene encoding sperm factor.

Indexing (details)


Subject
Cellular biology
Classification
0379: Cellular biology
Identifier / keyword
Biological sciences; Fertilization; Mammalian eggs; Oscillations; Sperm factor
Title
Characterization of the sperm factor responsible for initiating [calcium (2+)](i) oscillations during fertilization in mammalian eggs
Author
Wu, Hua
Number of pages
147
Publication year
2000
Degree date
2000
School code
0118
Source
DAI-B 61/10, Dissertation Abstracts International
Place of publication
Ann Arbor
Country of publication
United States
ISBN
0599964073, 9780599964075
Advisor
Fissore, Rafael A.
University/institution
University of Massachusetts Amherst
University location
United States -- Massachusetts
Degree
Ph.D.
Source type
Dissertations & Theses
Language
English
Document type
Dissertation/Thesis
Dissertation/thesis number
9988854
ProQuest document ID
304605860
Copyright
Database copyright ProQuest LLC; ProQuest does not claim copyright in the individual underlying works.
Document URL
http://search.proquest.com/docview/304605860
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