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INVESTIGATOR'S PROFILE

 



Prof.M. Hussain Munavar

 

 

 

 

 

 

 

                                                                        
Professor & Head
Department of Molecular Biology
Center for Excellence in Genomic Sciences
School of Biological Sciences
Madurai Kamaraj University, Madurai 625 021

Research Areas:

  • Genetics/Molecular Biology of Transcription control in Escherichia coli
  • Genetic Regulation by and of Proteases in Escherichia coli
  • Genetic regulation of DNA Repair and Mutagenesis in Escherichia coli 
  • Anticancer therapeutics and targeted drug delivery employing Escherichia coli minicells

Contact Details:
Email: munavar@rediffmail.com, munavar61@gmail.com
Phone/Fax : 0452-2458210

 

Research Interests:
Transcription control in E.coli:
On the area of transcription control, we have shown that the fitA and fitB genes earlier identified and very well characterized in E.coli in our laboratory are same as pheS and pheT genes.  This has lead to the new finding that the products of pheS and pheT   genes namely phenylalanyl tRNA synthetase also functions as a transcription factor.   By sequence analysis we have shown that the originally identified fitA76 mutant harbors two lesions:  a G293àA293 transition in pheS locus and another mutation, designated as  fit95.   Based on molecular analysis we have shown that fit95 defines an allele of the  pheT locus.   We have physically mapped the position of one of the earlier isolated fit mutation (fitC4), which was isolated as a suppressor of fitA76* mutant harboring pheS4 and fit95 mutations.  Our physical mapping indicate that fitC4  in all likelihood, could be duplicated copy of  a pheS4 mutation, which defines a GàC transversion at position 293 of pheS locus.  Also our biochemical analyses indicate that neither fit95, nor pheS5 mutations when present alone confer the phenotype characteristic of originally identified transcription mutant fitA76 (fit95 pheS5).  These results reinforce the conclusion that the phenylalanyl tRNA synthetase indeed functions as a transcription factor (Fit)  and two mutations are needed  to elicit the fitA76 Ts phenotype (pheS5 fit95). Moreover, we have clearly shown the fitC4 suppresses the Ts phenotype due to various fitA/B pheS/T mutations in an allele specific manner. Also, we have shown that the fit gene products do not function as global transcription factors but are involved in expression of only few classes of genes which might include genes for ribosomal proteins. During the course of the investigation we have embarked on to the identification of hitherto unidentified alternate promoter in the pheST operon of E. coli.
 Currently we are working in the following aspects:

  • Molecular characterization of the new alternate promoter that we have recently identified in pheST operon of E.coli and finding its  sigma dependence and Transcription Start Site.  Also we would try to identify transacting factors that potentially regulate the above alternate promoter.
  • Identification of  the lesion present in the fit95 mutant by PCR based amplification of fit95 region and  subsequent sequence analyses.
  • Isolation and characterization of promoter lacZ+ fusions which express beta galactosidase in Fit dependent manner and analyzing such promoter sequences in order to know whether Fit factors bind to specific sites on DNA to regulate transcription.
  • Molecular analysis of the transcription regulation by Cra

Proteolysis in E.coli:
My collaborative research work with Dr.Susan Gottesman of National Cancer Institute, National Institutes of Health, Bethseda, MD, USA, has lead to the new finding that to elicit the originally proposed Alternate Lon Protease, Alp+ phenotype in E. coli, atleast two proteases are needed.  The Alp protease originally was named so because in Alp+ strains the two substrates of Lon protease, namely SulA and RcsA, were expected to be degraded as like in a lon+ strain.  But, now it is shown that SulA is degraded by ClpYQ protease but the other function capable of RcsA inactivation/degradation is still not identified. 

Currently we are working in the following directions:
The faa mutation that we have identified and shown that it elicits Alp+ phenotype has been shown to be a mutation at position 232 of dnaJ gene of E.coli.  The Alp+ phenotype becomes stronger when ssrA mutation is also present.  The same Alp effect in a lon- strain is also seen in ssrA mutants which carry multicopy KanR plasmid.  In this area, we would like to do the following

  • Identifying and characterizing mutations that restore RcsA function in lon faa ssrA mutant of E.coli.  This will pave way to identify the actual gene coding for the function responsible for RcsA inactivation/decay. Later the activity can be demonstrated in vitro using purified  proteases, if possible. 
  • Identifying the other phenotypic traits affected by both the above said dnaJ mutation as well as the mutation that we anticipate to get.
  • Accidently it was found that lon mutants transformed with plasmid bearing truncated dnaK’ fragment showed Alp+ phenotype. The interesting molecular mechanisms behind this truncated dnak’ induced Alp/Lon Like protease activity, is being investigated currently. Intial mutational studies have given a clue that ClpYQ protease might be involved in RcsA degradation. Further studies may help to elucidate alternative proteolytic pathways for degradation of Lon substrates.

DNA Repair in E. coli:
An unorthodox DNA Repair which is SOS independent (named SIR) and specific to repair of Mitomycin C induced DNA damage was reported long ago from this Laboratory. This SOS Independent Repair was found to be elicited through a combination of Rif resistant allele named, rpoB87 and Nal resistant allele named, gyrA87 in recA defective strains or lexA3 Ind – strains. We revisited this issue to gain more understanding of this proposed SOS Independent DNA Repair (SIR) phenotype. PCR based cycle sequencing analysis revealed that rpoB87 is indeed defined by a mutation in the rpoB gene at the 522nd codon (TCT à TTT) located in cluster 1 of rifampicin resistance region of rpoB and gyrA87 is defined by a mutation in the 82nd codon of gyrA gene (GAC à AAC), a hitherto unreported allele of nalidixic acid resistance. Our studies show that these mutations together cause significant increase in resistance to Mitomycin C but not to UV as was expected. Our current efforts focus on

  • Unravelling the exact mechanism involved in the proposed SIR pathway and identifying the gene/genes taking part in the pathway.
  • Study of the activity of MMC repair specific genes like uvrB and demonstrating the involvement of even SOS genes in the pathway, if that were to be so.
  • Studying the effect of rpoB87 and gyrA87 mutations on general transcription and repression as rpoB87 allele is reported to be a fast moving RNA Polymerase and can increase transcription of SOS genes sans LexA repression and despite LexA repression. These studies may be extrapolated to orthodox lac and ara systems.

Mutagenesis:
Our recent work on informational suppression has led to the novel finding that the E. coli Amber suppressor, supE44, can also suppress Opal and Ochre mutations. Although the Wobble base pairing does not allow the Amber suppressor to suppress either Opal or Ochre, an alternative model for base pairing proposed by Lim and Curran lend some support to the above observation. During the course of the investigation on Post Plating Mutagenesis, a mutation isolated named ppm is now shown to be an allele of rpsD and it confers Ts phenotype on the cell. Sequence analysis reveals that ppm defines a single base deletion of T at the 570th base pair of the rpsD gene. In this aspect we would like to

  • Further characterize the ppm mutation and also isolate more ppm like mutations and characterize them
  • Studying the effect of other nonsense suppressors in the suppression of all types of Nonsense mutations
  • Perhaps embark on to study missense suppression and also study the effect of nusA mutations on Post Plating mutagenesis.

 
Recent Publications:

  1. Munavar, M.H. and Jayaraman, R.  “Extragenic suppression of the temperature sensitivity of a temperature sensitive transcription mutant of Escherichia coli”.  Presented a the 55th annual meeting of the Society of Biological Chemists held at Trivandrum, India (December 15-17, 1986)
  2. Munavar, M.H. and Jayaraman, R. Extragenic suppression of the temperature sensitivity of a fitA mutation by a fitB mutation in Escherichia coli: Possible interaction between fitA and fitB gene products in transcription control. J.Genet. 66:123-132, 1987
  3. Munavar, M.H.,  Madhavi, K. and Jayaraman, R.  Aberrant transcription in fit mutants of Escherichia coli and its alleviation by suppressor mutations. J. Biosci.  18:37-45, 1993
  4. Munavar, M.H. and Jayaraman, R.   Genetic evidence for the interaction between fitA, fitB and rpoB gene products and its implication in transcription control in Escherichia coli J. Genet. 72:21-33, 1993.
  5. Sankaran, S., Munavar, M.H. and Jayaraman, R.  “Phenylalanyl tRNA synthetase of Escherichia coli as an accessory transcription factor:  analysis of fitA76 and pheS5 mutations”.  Presented at the meeting on “Molecular Genetics of bacteria and Phages” held at Cold Spring Harbour Laboratory, CSH, NY, USA (August 24-29, 1993).
  6. Munavar, M.H. and Gottesman, S.  “Isolation and characterization of mutations affecting ALP  (Alternate Lon Protease) activity in Escherichia coli”.  Presented at the meeting on “Molecular Genetics of Bacteria and phages” held at the University of Wisconsin, Madison, WI, USA ( August  2-7, 1994).
  7. Munavar, M.H. and  Gottesman, S. “A chromosomal locus affecting Alternate lon-like protease (Alp)  activity in Escherichia coli”.  presented at the 95th General Meeting of the American Society for Microbiology held at Washington DC. (May 21-25, 1995).
  8. Ramalingam, S.,  Munavar, M.H., Sankaran, S.  Rukmani, A. and Jayaraman , R. Elucidation of lesions present in the transcription defective fitA76 mutant of  Escherichia coli:   Implication of phenylalanyl tRNA synthetase subunits as  transcription factors  J.Biosci. 1999  ,24;153-162
  9. Sudha, S. Munavar, M.H. and Jayaraman, R.  Synthesis versus stability of  RNA in fitA76 and pheS mutants of Escherichia coli  and its implications Indian J.Microbiol. 2001, 41:123-127.
  10. Hussain Munavar, YanNing Zhou. and  Susan Gottesman (2005), Analysis of Escherichia coli Alp phenotype: Heat shock induction in ssrA mutants:    J.Bacteriol.  187: 4739-4751
  11. Vidya.S., Praveen Kamalakar.B., Hussain Munavar M., Satish Kumar, L  and R. Jayaraman  (2006). Allele-specific suppression of temperature sensitivity of  fitA/ fitB mutants  of E.coli by a new  mutation (fitC4);  isolation, characterization its implications in transcription control.  J.Biosci. 31: 31-45
  12. B. Singaravelan, B. R. Roshini, and M. Hussain Munavar.(2010) Evidence that the supE44 Mutation of Escherichia coli Is an Amber Suppressor Allele of glnX and that It Also Suppresses Ochre and Opal Nonsense Mutations. J. Bacteriol. 192, 6039-6044
  13. R. T. Ponmani and M. Hussain Munavar. (2011) Assessing the Role of Cra in the Regulation of prpB and yahA genes of Escherichia coli in vivo using lacZ Transcriptional Fusions. Journal of Scientific and Industrial Research. (In Press).
  14. Thangaraj Ponmani and M. Hussain Munavar. (2010) Assessing the Role of Cra in the Regulation of prpB and yahA genes of Escherichia coli in vivo using lacZ Transcriptional Fusions. International Conference on Genomic Sciences held at SBS, MKU, Madurai (11th to 13th November, 2010) (Won the Best poster award)