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At high concentration, actinomycin D kills the protozon Chlamydomonas reinhardi with exponential kinetics. The rate of killing is dependent upon the temperature of incubation. This dependence is partially a function of the increased extent of binding of the drug at the higher temperature (33°C), but sensitivity of the cell must also be stimulated by the higher temperature. While actinomycin D probably kills cells in a reaction which requires binding to DNA, there is no correlation between the lethal event and the inhibition of macromolecular synthesis or the breakdown of macromolecules.

I have developed a simple model to explain the difference in sensitivity of various species of RNA to actinomycin D inhibition. This model predicts that frequently transcribed genes will be much more sensitive to the drug than infrequently transcribed genes.

Mutants of Chlamydomonas reinhardi have also been isolated which are both temperature sensitive in growth and resistant to kill­ing by actinomycin D. These mutants, unlike other actinomycin D­ resistant cell lines, are neither impermeable to the drug nor do they excrete it at an accelerated rate. The mutants are partially temper­ ature sensitive in their ability to synthesize RNA. In the presence of actinomycin D, however, RNA synthesis is partially protected at the nonpermissive temperature (and, in some cases, at the permissive temperature also) when compared to the inhibition of wild type cells.

Extraction and examination of RNA from these mutants reveals that actinomycin D inhibits different species of RNA to different extents.

I propose that the mutants have an altered chromosomal constituent, which impedes the binding to the genome. At the nonpermissive temperature the alteration is postulated to partially interfere with transcription.

In the course of these experiments it became necessary to determine the maturation pathways of the ribosomal RNA species of Chlamydomonas reinhardi. Cytoplasmic rRNAs of C. reinhardi are cleaved from a single precursor of molecular weights 2.4 · 106 to a mature rRNA (0.69 · 106mol.wt) and a 1.4 · 106-mol. wt precursor of a mature 1.3 · 106-mol. wt rRNA. The kinetics of incorporation of radioactive label into the rRNAs suggest that the 0.69 · 106-mol. wt rRNA gene is located closer to the promotor than is the gene for the 1.4 · 106-mol. wt rRNA. The synthesis of cytoplasma rRNAs is extremely sensitive to camptothecin, an inhibitor of nuclear rRNA synthesis, but synthesis of chloroplast rRNA is quite resistant to the inhibitor. This has allowed us to demonstrate that chloroplast rRNAs are processed from precursors which resemble those of blue-green algae. A 1.14 · 106-mol. wt precursor is processed to the 1.07 · 106-mol. wt mature chloroplast rRNA, and a 0.64 · 106-mol. wt precursor is cleaved to a 0.56 · 106-mol. wt species and then to the mature 0.54 · 106-mol. wt rRNA. This study demonstrates two new ways in which the function of the chloroplast genome resembles those of prokaryotes more than those of the nucleus.


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CaltechTHESIS  

Autor(es)

Miller, Mark James - 

Id.: 70077808

Versión: 1.0

Estado: Final

Tipo:  application/pdf - 

Tipo de recurso: Thesis  -  NonPeerReviewed  - 

Tipo de Interactividad: Expositivo

Nivel de Interactividad: muy bajo

Audiencia: Estudiante  -  Profesor  -  Autor  - 

Estructura: Atomic

Coste: no

Copyright: sí

Formatos:  application/pdf - 

Requerimientos técnicos:  Browser: Any - 

Relación: [References] http://resolver.caltech.edu/CaltechTHESIS:09072017-112605896
[References] http://thesis.library.caltech.edu/10418/

Fecha de contribución: 08-sep-2017

Contacto:

Localización:
* Miller, Mark James (1976) Investigations of the Mechanism of Cell Killing Induced by Actinomycin D. Dissertation (Ph.D.), California Institute of Technology. http://resolver.caltech.edu/CaltechTHESIS:09072017-112605896

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