How cAMP Homeostasis is Controlled in Leishmania by Receptor Adenylate Cyclases and Acidocalcisomal Pyrophosphatases Aiding its Survival in Phagolysosomal Conditions?

Abstract

Leishmania donovani while infecting macrophages is exposed to phagolysosome conditions (PC) (37 °C and pH 5.5) from the sandfly gut environment of 22 °C and pH 7.4. This shift in temperature and pH led to increased intracellular cAMP and cAMP‐mediated PKA activity in the parasites, which help in intra‐macrophage survival, a pre‐requisite for infectivity and transformation to amastigotes. Though cAMP and cAMP‐triggered signaling pathways seemed to be one of the important factors for parasite survival and infectivity, the mechanism of cAMP production and its stringent control is obscure. In the absence of typical orthologs for G‐proteins and G‐protein coupled receptors, the precise mechanisms for positive modulation of cAMP production during exposure to PC seemed an important aspect to be studied. Amongst various membrane bound isoforms of receptor adenylate cyclase, two promastigote‐stage specific membrane bound receptor adenylate cyclases (LdRAC‐A and LdRAC‐B), LdRAC‐A appeared to function as a major cAMP generator following PC exposure. Following PC exposure, pyrophosphate (PPi), an energy storage compound as well as a by‐product of cAMP biosynthesis by adenylate cyclase, was found to be decreased. This may be due to microtubule and microfilament‐driven translocation of acidocalcisomes near plasma membrane vicinity with concomitant increase of acidocalcisome membrane pyrophosphatase (LdV‐H+PPase) and acidocalcisomal soluble pyrophosphatase (LdVSP1). Episomal over‐expression and conditional silencing demonstrated regulatory role of V‐H+PPase on cAMP trigger and consequent induction of resistance to macrophage‐derived pro‐oxidants and parasite killing. Moreover, immunofluorescence analysis indicated towards a possible colocalization of LdV‐H+PPase and LdRAC‐A during PC exposure. The results suggested that translocation of acidocalcisome in membrane vicinity functions as a trigger for LdRAC‐A‐driven cAMP generation through depletion of PPi pool by LdV‐H+PPase. For the first time, one of the molecular mechanisms behind the positive modulation of cAMP by leishmanial adenylate cyclases in absence of Gproteins was deciphered by us.Support or Funding InformationDST‐INSPIRE, Department of Science and Technology (DST), Govt of India, DST‐PURSE, Department of Science and Technology (DST), Govt of India, UGC‐SAP, UGC, Govt of IndiaThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.