A. Selvapandiyan1 , P. Kumar2 , J. C. Morris3 , C. C. Wang2 , H. L. Nakhasi1 , 1CBER, FDA, Bethesda, MD, 2UCSF, San Francisco, CA, 3Clemson University, Clemson, SC
Background: Leishmania donovani, is a causative agent of fatal Visceral disease in humans. At present there is no vaccine for such disease. To develop an effective vaccine, one needs to know the pathogenesis of this parasite. We have characterized several genes which play important roles in parasite growth. One such gene is centrin. Recently, we identified several isoforms of this gene in Leishmania. In the present study we have undertaken to analyze the function of centrin isoforms in a related trypanosomatid, T. brucei using RNA interference (RNAi) methodology.
Methods: RNAi is a mechanism, where introduction of homologous double stranded RNA specifically targets a gene product to make a null phenotype.
Results: Depletion of all the five centrin gene products by RNAi in the procyclic form of T. brucei, revealed, that out of five centrins, only the knockdown of centrins 1, 2 and 3 affected the growth of the parasite in vitro. Microscopic observation revealed that these centrin deficient procyclic cells became enlarged in size and contained multiple basal bodies, kinetoplasts, Golgi, nuclei and detached flagella. These multiple organelles were, however, closely clustered together, indicating duplication without segregation in the absence of centrins. This failure in organelle segregation constitutes the likely cause of cytokinesis inhibition, which in turn results in growth inhibition.
Conclusions: This study suggests a unique role for centrin in the segregation of multiple organelles in the procyclic-form of T. brucei and therefore plays an important role in parasite cell division and growth.This study further demonstrates the importance of centrin genes in trypanosomatid pathogenesis.