Centre for Molecular and Cellular Biology
Current projects
Tuberculosis
TB research forms the largest component of work in the centre,
but may be subdivided into a number of smaller projects.
Molecular
Epidemiology
M. tuberculosis
strains from patients in two local suburbs are genetically fingerprinted using
techniques based on the transposition element, IS6110, and other probes. Together
with clinical data, these fingerprints are then used to study the dynamics
of the epidemic.
Different strain types
identified by this technique have been shown to have different rates of transmission
and levels of representation in the community. We have calculated transmission
rates to be surprisingly low, indicating that disease due to reactivation
of supposedly latent strains, also drives the epidemic.
We are also investigating
the mutation rate of the fingerprint patterns, which appear to be considerably
lower than those observed in studies from other parts of the world.
The emergence of drug-resistant
strains of Mycobacterium tuberculosis, especially multidrug-resistant (MDR)
strains, poses a threat to the success of tuberculosis control programs and
places an enormous financial burden on the health budget.
This project focuses on
understanding drug resistance in local communities and our recent results
can be summarised as follows:
- Drug resistance is
due to mutations in certain genes.
- About 60% of drug
resistance in local communities is due to transmission from person to person
and this indicates limitations in the control program.
- A molecular based
dot blot hybridisation method has been developed for quick screening of
drug resistance. This method also forms the basis of molecular technology
transfer to different countries in Africa.
- Micro-epidemics of
drug resistant strains have been identified in local communities
Currently the project
aims to establish a study site in order to investigate the clinical usefulness
of molecular prediction of drug resistance for identifying patients infected
with drug resistant M. tuberculosis as an adjunct to the local TB control
program. We hope this research will lead to the implementation of some community
intervention activities in the future.
Genomics
The correlation
of genetic and phenotypic polymorphism in M. tuberculosis is another aspect
of the TB project. We have focused on the impact of transposition of the insertion
element IS6110 into various locations in the bacterial genome. Our studies
have shown that about b of insertions occur within coding regions.
We have also investigated
various aspects of the large PPE gene family, including its polymorphic nature,
as well as the subcellular location of PPE proteins. Our data demonstrates
that the representative protein used is hydrophobic and cell-wall associated.
Currently, our main focus
is to determine the transcription start sites of some PPEs ,which will enable
us to compare the promoter regions of other PPEs in order to understand the
factors controlling their expression.
The M. tuberculosis genome
contains a family of putative subtilisin-like Serine Protease genes, termed
Mycosins, the function of which is unknown. Current research is directed at
investigating the significance of members of this seemingly abundant family.
Our findings so far have
lead us to hypothesise that the proteins encoded by the gene clusters including
the mycosins are involved in the secretion, processing, and activation of
the immunologically important ESAT-6 and CFP-10 proteins.
The main focus of our
efforts is directed at understanding the role of the subtilisin-like serine
proteases in the cleavage and activation of these immunologically important
peptides after secretion.
Immunology
An aspect
of our research involves the delineation of the host humoral (antibody) response
toward M. tuberculosis antigens which, it is hypothesised, is expressed differentially
during treatment. Differential expression as treatment progresses can be tracked
as a predictor of treatment outcome. We hope to understand why some strains
are more virulent than others, as well as why the disease persists in some
patients.
Susceptibility to M. tuberculosis
may be influenced by environmental and/or genetic factors, with both potentially
affecting the immune response. A type 1 immune response is required for the
elimination of Mtb, while a type 2 immune response is essential for the eradication
of intestinal parasites.
Given that type 2 cytokines
inhibit type 1 responses, we hypothesise that susceptibility to TB in the
local community may be increased in individuals mounting prominent type 2
responses.
We have found increased
parasite infection and serum IgE levels in areas with the highest TB incidence.
We have also found evidence that BCG immunisation may protect children against
parasite infestation.
The complex network of
cytokine, chemokine and receptor expression in the immune system response
to M. tuberculosis forms the focus of a project investigating the heterogeneous
protein expression profiles within patients during their immune response to
M. tuberculosis by in situ hybridisation at infection site.
We are also investigating
whether differences exist between the adult and childhood immune responses
to M. tuberculosis infection. We anticipate that these studies will shed a
greater understanding on the heterogeneity of the host response.
Host
Genetics
Even
though roughly one third of the world's population is infected with M. tuberculosis,
the vast majority of these persons will never develop any clinical disease.
Inter-individual variation in the immune response against M. tuberculosis
plays a major role in determining the different clinical manifestations and
outcomes in infected persons. Our study is to identify some of these unknown
host genetic factors influencing immunity. This will provide a better understanding
of the pathogenesis of tuberculosis in humans, enabling better and novel approaches
to prevention and therapy.
Surrogate
Markers
This
project aims to find a set of markers, be they clinical, immunological or
bacteriological, that can be used to predict which patients will respond poorly,
or even have a second infection (relapse), after completion of TB therapy.
Here the role is to look at mycobacterial factors that might contribute to
persistence of disease and/or treatment failure.
Pharmacology
Arylamine
N-acetyltransferase 2 (NAT2) is the Phase-II metabolising enzyme in the body,
responsible for the inactivation of isoniazid, the mainstay anti-tuberculosis
drug. Phenotypic analyses of the activity of NAT2 have shown that the rate
of acetylation varies between individuals as well as in various ethnic populations.
These phenotypic changes can be correlated with well described genetic mutations
in the gene. We hypothesise that NAT2 polymorphism of the individual may influence
the efficacy of their treatment, and subsequently, even the possible epidemiology
of TB within the local population Thus far, we have shown that the NAT2 allele
distribution in patients shows a major shift from that seen in normal unaffected
individuals.
Genetics
of Cardiovascular Diseases
Hypertrophic
Cardiomyopathy
HCM is
an inherited form of heart disease, being the number one killer of sports
people under 35, and often the cause of youngsters dying suddenly on the rugby
field. We have been actively involved in defining the mutations that cause
the disease in SA. From a global perspective, it is now known that HCM is
caused by more than 100 different defects in the proteins of the contractile
apparatus of the heart muscle. Some of these proteins, like myosin and actin,
have been well studied.
We are particularly interested
in studying Myosin Binding Protein C (MyBPC). The exact function of MyBPC
is unknown, yet we know that it must fulfil an important role, because mutations
in certain domains cause HCM. The nature of this function is the main thrust
of our research. Some successes in our work thus far have lead to a fruitful
collaboration with Prof Hugh Watkins from Oxford University.
Progressive
Familial Heartblock Type II (PFHB-II)
The initial
aim of this study was to map PFHB-II to a chromosomal locus. This was achieved
by screening family members in which the disease occurs, with linkage analysis
of genetic markers at candidate loci.
This process of linkage
analysis gave us a statistical probability score that the PFHB-II-causative
gene was located on chromosome 1q32.-1q41. Our findings subsequently led us
to the actual search for the PFHB-II gene.
In silico research, using
data from the Human Genome Project has allowed us to identify certain genes,
as well as novel, previously uncharacterised genes -"virtual genes"
- as viable candidates for PFHB-II. This has reduced the time taken to identify
PFHBII-candidate genes. These new candidate genes are now being screened for
mutations that may cause PFHBII.
Psychiatric Disorders
Obsessive-compulsive
disorder
Obsessive-compulsive
disorder (OCD) is a disabling psychiatric disorder, with an underlying genetic
component. Pharmacological studies of its pathophysiology implicatesthe involvement
of members of the serotonergic and dopaminergic systems in the body. Using
case-controlled association studies of candidate genes in these two systems,
it may be possible to create a profile of biological markers, which would
allow for better diagnosis, as well as defining a treatment which is more
individually-specific. Research into the field of psychiatric genetics is
still in infancy, and finding genes, which confer susceptibility to OCD (and
other psychiatric disorders), is still a long way away. However, preliminary
data generated by our group, and many others around the world, looks very
promising in achieving the aim of a genetic based diagnostic regimen.
|
