Student Theses and Dissertations

Date of Award

2007

Document Type

Thesis

Degree Name

Doctor of Philosophy (PhD)

RU Laboratory

Shaham Laboratory

Keywords

non-apoptotic, programmed cell death, linker cell, C. elegans, caspase-independent, developmental timing

Abstract

The most intensively studied form of programmed cell death (PCD) is apoptosis, which is characterized by stereotypical morphological features including chromatin compaction and by a requirement for the activity of caspase proteases, which are controlled by conserved gene pathways. Although non-apoptotic, caspase-independent programmed cell death pathways have been postulated, there is little evidence to convincingly prove their existence, and few insights regarding their molecular basis or possible in vivo functions. To investigate this question, we have studied the developmentally regulated PCD of the Caenorhabditis elegans linker cell. We have carried out transmission electron microscopy studies of dying linker cells, which revealed non-apoptotic features, including nuclear crenellation in the absence of chromatin condensation, swelling of mitochondria and endoplasmic reticulum, and accumulation of cytoplasmic single- and multilayered membrane-bound structures. Similar morphological changes occur during the normal developmental death of some vertebrate neurons in the spinal cord and ciliary ganglia, suggesting that this is a highly conserved cell death program. Our genetic studies demonstrate that linker cell death is a non-apoptotic programmed cell death. This cell death is independent of the ced-3 caspase, other C. elegans caspase homologs, and can occur even when a broad-spectrum caspase inhibitor is expressed. We have found that the engulfment of the linker cell is independent of the known C. elegans engulfment genes. We tested and found no evidence for the involvement of autophagic, necrotic, or Wallerian degeneration genes in linker cell death. By ablating cells neighboring the linker cell, and by examining mutants in which the linker cell is abnormally positioned, we demonstrated that the linker cell employs a cell-autonomous program to promote its demise. Using a candidate gene approach, we showed that linker cell death is controlled by the microRNA let-7 and by the zinc finger transcription factor lin-29, both components of the main developmental timing pathway in the animal. Conducting a genome-wide RNAi screen, we have identified new candidate regulators of linker cell death. Characterization of these genes may uncover the molecular mechanism driving this new type of programmed cell death in C. elegans.

Comments

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

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