Student Theses and Dissertations

Date of Award

1991

Document Type

Thesis

Degree Name

Doctor of Philosophy (PhD)

RU Laboratory

Young Laboratory

Abstract

The period (per) gene of Drosophila melanogaster is fundamentally involved in the generation of biological rhythms. Three classes of per mutations which alter circadian periodicity have been identified: pers mutants have circadian behavioral rhythms of 19h instead of 24h; per mutants have long period rhythms of 28h; and per0 mutants have no detectable circadian rhythms. Steps have been taken to gather more information about per's role in the construction or maintenance of biological clocks. By analyzing transformed Drosophila lines, the amount of per product was found to be integral to the pace of the clock. Absence of. the per product leads to arrhythmicity; more per product shortens the period length; less per product lengthens period. In addition, single amino acid changes in the per product can mimic these results. DNA sequence analysis has revealed that in per0 flies, a single nucleotide change resulted in a translational stop codon and hence a truncated protein. A valine-to-aspartic acid change in the per1 mutants lengthens period. Likewise the shortened period length in pers mutants is a result of a serine-to-asparagine substitution. These combined studies suggest that per1 and pers mutants produce hypoactive and hyperactive per proteins, respectively. Using the sequence analysis of the mutants as a starting point, further amino acid changes in per were created, introduced back into the fly, and then evaluated for effects on biological rhythms. Five out of six amino acid changes near the pers mutation also gave short period lengths. These results suggest that the region near the permutation acts as a domain to restrain per function and thereby slows the clock. Further insight into the nature of per function was obtained through a cell level assay. The per mutations have a significant effect on intercellular communication in the salivary gland cells of third instar larvae. Dye transfer and electrophysiological experiments indicate that gap junction conductances varies inversely with the period of the behavioral rhythms. Such alterations in communication in the nervous system may explain how per influences biological rhythms. Lastly, a detailed localization study of the per gene products during embryogenesis shows that it is expressed in particular cells in the brain and ventral nerve cord. This information should make it possible to localize the focus of per's clock function to specific cells.

Comments

A thesis submitted to the faculty of the Rockefeller University in partial fulfillment of the requirements for the degree of Doctor of Philosophy

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