Molecular Basis of a Digital, Multi-state Developmental Switch

University Of California-Santa Barbara, Santa Barbara CA

Investigators

Abstract

Intellectual Merit. Stem cells of multicellular organisms differentiate into diverse specialized cell types through the action of multi-state developmental switches. The ultimate aim of this project is to understand the molecular mechanism of mating type determination in Tetrahymena thermophila, as a model of a multi-state developmental switch. Ciliated Protozoa (including T. thermophila) are unicellular eukaryotes that maintain two copies of their genome. Seven distinct mating types ("genders") occur in T. thermophila, numbered I to VII. In order to mate (conjugate), physical contacts between cells of different mating type are required. The micronucleus contains the genetic information for five to seven mating types, but only one mating type is chosen to be expressed from the macronucleus. The choice of which mating type will be expressed takes place during differentiation of the macronucleus. The mating type to be expressed is chosen randomly, as if a roulette wheel were making the choice. The choice, once made, is irreversible. Thus the mat locus thus behaves as multi-state, developmental switch, which is reliably, permanently, and randomly set during MAC differentiation to one somatically heritable state. This project aims to test the following working hypothesis. During macronuclear differentiation, mating type determination involves a progressive wave of chromatin remodeling moving through the mating type locus, believed to contain up to seven distinct mating type genes in a row. The wave stops at a random location in the mating type locus and the DNA of remodeled mating type genes is deleted. The first undeleted gene becomes the expressed mating type gene. The results of this project are expected to contribute knowledge about the molecular basis of multi-state developmental switches. The results are also expected to extend knowledge in the area of chromatin remodeling guided by small interfering (regulatory) RNAs. Because the hypothesized mechanisms are evolutionarily broadly conserved among eukaryotes, the project may well contribute insights into developmental switches of general significance, e.g., the switch that patterns differentiation along the anterior-posterior axis during embryonic development of mammals and other metazoa, or switches that determine the huge diversity of immune system cells that recognize and destroy microbial invaders of the vertebrate circulatory system. Broader Impacts. This project is expected to generate important new knowledge, genomic and gene sequence data and novel constructs that should be a boon to Tetrahymena molecular genetics and enhance the value and contributions of this valuable model organism for fundamental and applied research - as reflected by the 2009 Nobel Prize in Medicine. New genetic and genomic resources will be deposited in public databases and the Tetrahymena Stock Center. The project will additionally invest in the future by training a postdoctoral scholar. In addition, a group of 20-30 talented undergraduates will be involved in carrying out original research (for an average period of one year) that contributes to practically every aspect of the project. Historically, a large and highly diverse group of undergraduates has done original research in the principal investigator's lab (including high proportions of women and members of under-represented minorities); this will continue to be encouraged.