Date of Award
Fall 1996
Document Type
Dissertation - Restricted
Degree Name
Doctor of Philosophy (PhD)
Department
Biological Sciences
First Advisor
Waring, Gail L.
Second Advisor
Fredericks, Walter W.
Third Advisor
Karrer, Kathleen M.
Abstract
Understanding the molecular mechanisms that underlie generation of complex three-dimensional structures is one of the most fascinating and challenging problems in developmental biology. The Drosophila eggshell formation represents a favorable model to study the assembly of suprarnolecular structure during development in a genetically amenable eukaryote. Among the various eggshell genes cloned, dec-1 (defective chorion) offers a good opportunity to analyze the role of proteolysis in the in vivo regulation of an assembly pathway.· The dec-1 locus generates three proteins of 106 kDa (fcl06), 125 kDa (fcl25), and 177 kDa (fcl77). Previous studies established that fcl06 is first processed into a protein of about 80 kDa, which in turn, is cleaved into a c-terminal derivative of approximately 60 kDa (s60). Preliminary observations indicated that fcl25 may also be processed during oogenesis. Polyclonal antibodies produced against trpE-fusion proteins containing selected regions of dec-1 open reading frame were used to further analyze the proteolytic processing of .the dec-1 proteins as well as their in situ distribution in the eggshell. It was found that fcl06, fcl25 and fcl77 follow different regulated maturation pathways that result in the production of several stable proteins with distinctive N- and C-termini. Each one of the dec-1 mature proteins shows a different distribution in the eggshell. fcl06 processing generates three final products, the previously identified s60, a 25 kDa (s25) and a 20 kDa (s20) proteins. s60, fractionates between the vitelline membrane (VM) and the chorion, the two main layers of the eggshell; s25 is a chorion component while s20 localizes into vesicles inside the oocyte. fc125 processing yields a stable protein of approximately 95 kDa (s95) which during early choriogenesis is observed in the VM and in the forming chorion. The final processing derivative of fc177 localizes in the mature eggshell, inside rounded cavities characteristic of the shell chorion. The different behavior in the egg chamber of the various mature dec-1 proteins indicates that there is no functional redundancy among them. The complexity of the dec-1 locus suggests that the assembly of the eggshell is a highly regulated process.