Genomic analysis is a new approach for the characterization and
investigation of novel genes, gene clusters, the function of uncharacterized
proteins, and genetic diversity in microorganisms. These approaches are important
for the study of chlamydiae, a system in which several genomes have been
sequenced but in which techniques for genetic manipulation are not available. The
objective of this thesis is to combine computer-based analysis of chiamydial
inclusion membrane proteins (Incs) with cellular and molecular biological analysis
of the bacteria. Three different experimental lines of investigation were examined,
focusing on Incs of C. trachomatis and C. pneumoniae.
Chlamydiae are obligate intracellular bacteria that develop within a nonacidified
membrane bound vacuole termed an inclusion. Putative Inc proteins of C.
trachomatis and C. pneumoniae were identified from genomic analysis and a
unique structural motif. Selected putative Inc proteins are shown to localize to the
inclusion membrane.
Chiamydia trachomatis variants with unusual multiple-lobed, nonfusogenic,
inclusion were identified from a large scale serotyping study.
Fluorescence microscopy showed that IncA, a chiamydial protein localized to the
inclusion membrane, was undetectable on non-fusogenic inclusions of these
variants. Sequence analysis of incA from non-fusogenic variant isolates revealed a
defective incA in most of the variants. Some variants lack not only IncA on the
inclusion membrane but also CT223p, an additional Inc protein. However, no
correlation between the absence of CT223p and distinctive inclusion phenotype
was identified. Nucleotide sequence analysis revealed sequence variations of C.
trachomatis incA and CT223 in some variant and wild type isolates.
Comparative analyses of the three recently published C. pneumoniae
genomes have led to the identification of a novel gene cluster named the CPn1O54
gene family. Each member of this family encodes a polypeptide with a hydrophobic
domain characteristic of proteins localized to the inclusion membrane. These
studies provided evidence that gene variation might occur within this single
collection of paralogous genes. Collectively, the variability within this gene family
may modulate either phase or antigenic variation, and subsequent physiologic
diversity, within a C. pneumoniae population.
These studies demonstrate the genetic diversity of Inc proteins and
candidate Inc proteins, within and among the different chiamydial species. This
work sets the stage for further investigations of the structure and function of this set
of proteins that are likely critical to chlamydial intracellular growth. / Graduation date: 2003
| Identifer | oai:union.ndltd.org:ORGSU/oai:ir.library.oregonstate.edu:1957/31603 |
| Date | 06 June 2002 |
| Creators | Viratyosin, Wasna |
| Contributors | Rockey, Daniel D. |
| Source Sets | Oregon State University |
| Language | en_US |
| Detected Language | English |
| Type | Thesis/Dissertation |
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