Identification and characterization of a novel component of the contractile ring
Abstract
In order to identify and characterize novel components of the contractile ring, a monoclonal antibody library was prepared against a mixture of actin-associated proteins from sea urchin eggs. These antibodies were screened for positive staining of the contractile ring in egg cortices isolated at the time of cytokinesis. The antigen recognized by monoclonal antibody 68-13 was enriched in the contractile ring relative to the adjacent cortex. In immunoblots, this antibody recognized two high molecular weight proteins in sea urchin eggs and sea cucumber smooth muscle. The smooth muscle was used to purify the 68-13 antigen. A 900 kD protein was isolated and polyclonal antibodies were made against this protein. The 900 kD fractions also contained a 300 kD proteolytic fragment. The 900 kD and the 300 kD bands were stained by both the monoclonal and polyclonal 68-13 antibodies The term 68-13 antigen refers to both the 900 kD and the 300 kD bands. The polyclonal antibody stained the contractile ring. The 68-13 antigen bound myosin in blot overlay assays. Sequence information from proteolytic fragments of the 900 kD protein indicated that this protein is a titin-like molecule. Localization studies indicated that titin, actin, and myosin were present at the same time in the contractile ring of dividing sea urchin eggs.
Degree
Ph.D.
Advisors
Otto, Purdue University.
Subject Area
Cellular biology|Molecular biology
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