Epistatic Analysis of Genes Involved In Streptococcus mutans Mutacin Production

Objectives: Streptococcus mutans are capable of out-competing other gram-positive oral flora due of its ability to produce a lantibiotic Mutacin I. Previous studies identified 44 genes/operons involved in the production of mutacin I. To further analyze the regulatory network of mutacin I production, Epistatic Miniarray Profile (E-MAP), was adapted to categorize genes involved in mutacin I production.

Method: Both single and pair-wise mutations were constructed for genes selected in this study. Analysis was performed by measuring both mutacin I production and mutA promoter activity of all mutants compared to wild-type. LOESS regression was performed to obtain predicted behaviors for all pair-wise mutations and an interaction score were then calculated for every gene pair. The level of interaction between any two genes in this study were then determined using hierarchical clustering of the interaction scores.

Results: In this pilot study involving 8 genes. Smu1850 (pepO) and Smu1034 (pepN) were clustered together, indicating a strong interaction between these two genes. Both genes encode zinc metalloendopeptidases involved in protein degradation and mutation of both genes enhanced mutacin prodiction as well as mutA gene transcription. Smu1311, an UDP-N-acetylglucosamine 2-epimerase was found to interact strongly with ciaH, a histidine kinase involved in regulating mutacin production.

Conclusion: Results from our pilot E-MAP study demonstrated the feasibility of applying this technique in revealing important relationships among genes involved in mutacin production. One limitation of this technique is in predicting interaction among regulatory genes. However, the accuracy of the prediction increase in correlation with the number of genes involved in the analysis, therefore we expect a greatly improved and comprehensive results when all 44 genes/operons from our library screening are analyzed. Results obtained from E-MAP analysis can be further complemented with classical epistatic analysis as well as providing candidates for microarray analysis.

Grant: NIH R01DED14757