Thesis (M.S.)--University of North Texas, Aug., 2007. / Title from title page display. Includes bibliographical references.

No description available.

published_or_final_version / Paediatrics and Adolescent Medicine / Doctoral / Doctor of Philosophy

Thesis (M. Sc. (Chemistry) -- University of Pretoria. 2006. / Includes bibliographical references (leaves 111-117).

No description available.

Mycobacterium tuberculosis (MTB) encodes three enzymes that are dependent on vitamin B12–derived cofactors for activity, including a B12-dependent methionine synthase (MetH). Previously, work in the Molecular Mycobacteriology Research Unit (MMRU) demonstrated vitamin B12 auxotrophy in a mutant strain disrupted in the alternative, B12-independent methionine synthase, MetE. This observation established the ability of MTB to transport corrinoids despite the absence of an identifiable B12-specific transporter. In addition, it suggested that MTB does not synthesize vitamin B12 in vitro. Notably, bioinformatic analyses identified PPE2 as the only B12-related transport candidate in MTB, though as a putative B12-regulated cobalt transporter. PPE2 is unusual in possessing directly upstream of its predicted start codon one of only two B12-dependent riboswitches in the MTB genome, and it lies in a putative operon with B12 biosynthetic genes, cobU and cobQ1. In this study, the possibility that PPE2 functions in the transport of vitamin B12 or cobalt was investigated. Transcriptional and phenotypic data suggested that PPE2 was not involved in B12 transport. Instead, it was shown that cobalt can supplement the growth of an MTB metE mutant in liquid medium, strongly supporting the ability of MTB to synthesize B12 de novo. Moreover, the ability to utilise exogenous cobalt was dependent on functional PPE2, thereby establishing a role for a PPE-family member in cobalt assimilation in MTB. Vitamin B12 comprises a central corrin ring co-ordinated to 5,6-dimethylbenzimidazole (DMB) as α-axial ligand. Substituting DMB with adenine yields the alternate form, pseudo-B12. The ability of mycobacteria to utilize pseudo-B12 precursors (cobinamide and adenine) to support full function of B12-dependent metabolic pathways was evaluated. Although the pseudo-B12 precursors appeared to complement chemically the mycobacterial B12 auxotrophs, growth of the mutants on cobinamide alone complicated this interpretation. To address this limitation, DMB synthesis was targeted by disrupting the MTB bluB homologue, Rv0306. Neither site-directed mutagenesis of key Rv0306 residues, nor full-gene deletion was sufficient to eliminate growth on cobinamide. Instead, this observation highlights the need to establish biochemically the nature of the active B12 form synthesized and utilized by MTB under different conditions. In combination, the results presented here support the inferred flexibility of vitamin B12 biosynthesis in MTB, and reinforce the potential role of B12-dependent metabolism in mycobacterial pathogenesis.

Acquired Immunodeficiency Syndrome (AIDS) and tuberculosis (TB) have posed diagnostic and therapeutic challenges globally. Nowadays, it is estimated that 34 millions people are living with Human Immunodeficiency Virus (HIV). About 2 millions of people die from AIDS-related causes currently in each year. Tuberculosis is the most common presenting illness and leading cause of death among AIDS patients. Emerging studies suggest that HIV and Mycobacterium tuberculosis (Mtb), the causative pathogen of TB, act synergistically to accelerate decline of immune functions and cause the death. Mtb infection usually remains latent. Only small portion of infected individuals develops active TB. However HIV infection boosts the risk of reactivation of TB and susceptibility to new Mtb infection. In contrast, Mtb infection dysregulates cytokines production and induces HIV viral replication. Although it is well-known that HIV and Mtb potentiate each other in disease development, mechanisms of interaction of the two pathogens remain not well-elucidated. The aim of this study is to investigate the interaction of HIV viral protein Tat with mycobacteria infection, which may provide insights in the interplay between HIV and Mtb infections. HIV viral transactivator protein, Tat, plays a critical role in HIV replication; and its induction of apoptosis in CD4+ T cells contributes to immune defects. In this study, Tat was demonstrated to dysregulate immune responses against mycobacteria such as autophagy, a tightly regulated bacterial clearance mechanism. With pretreatment of the primary human blood monocyte-derived macrophages with Tat, the interferon-γ (IFN-γ)-induced Signal Transducer and Activator of Transcription-1 (STAT-1) phosphorylation was suppressed. Inhibition of STAT-1 phosphorylation ultimately led to downregulation of autophagy-associated gene, microtubule-associated protein light chain 3 (LC3) expressions. Of note, Tat was demonstrated to inhibit the colocalization of Bacillus Calmette Guerin (BCG) and IFN-γ-induced autophagosomes under fluorescent microscopy examination. In addition to the inhibition of bactericidal autophagy, Tat was found to manipulate cytokines production. Tat was demonstrated to enhance mycobacteria-induced tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) production. TNF-α and IL-1β have been well-demonstrated in literatures that can limit bacterial growth. They, however, have been also shown as important contributors to the increase of HIV viral replication in HIV and mycobacteria coinfection. Mtb-induced TNF-α production can induce transcriptional activation of the HIV long terminal repeat (LTR) promoter while blocking of IL-1β production decreases HIV replication. Tat enhancement of these cytokines production may therefore contribute to the knowledge of the increased viral replication in HIV and mycobacteria coinfection. Furthermore, new microRNAs, up-and-coming fine-tuners of innate immunity, were discovered. MicroRNAs, a family of non-coding RNAs, can regulate gene expressions post-transcriptionally and control various developmental and cellular processes. They can target mRNAs of cellular signaling molecules, transcription factors or cytokines as to regulate the immunity. Herein, microRNA-1303, originally with unknown function, was shown to regulate mycobacteria-induced TNF-α production and affect the Tat enhancement of TNF-α production. Taken together, the results of this study demonstrated that HIV viral protein, Tat could dysregulate immune responses to mycobacteria. The study of the dysregulation may further elucidate the interplay between HIV and mycobacteria infections. / published_or_final_version / Paediatrics and Adolescent Medicine / Doctoral / Doctor of Philosophy

Thesis (Ph. D.)--University of Hong Kong, 2008. / Also available in print.

Thesis (Ph. D.)--University of Hong Kong, 2008. / Also available online.

Thesis (Ph. D.)--University of Missouri--Columbia, 1996. / "August 1996" Typescript. Vita. Includes bibliographical references. Also available on the Internet.