Calcimycin mediates mycobacterial killing by inducing intracellular calcium-regulated autophagy in a P2RX7 dependent manner

Abstract

AIM - To understand the mechanism of autophagy induction by calcimycin and its effect on the viability of intracellular mycobacteria in THP-1 cells. RESULT - We noticed that treatment with calcimycin led to up-regulation of different autophagy markers like Beclin-1, autophagy-related gene (Atg) 7, Atg 3 and enhanced microtubule-associated protein 1A/1B-light chain 3-I (LC3-I) to LC3-II conversion in macrophages. This calcimycin-mediated killing of intracellular M. smegmatis and M. bovis BCG was abrogated in the presence of 3-methyladenine (3-MA). We also demonstrate that calcimycin binding with purinergic receptor P2X7 (P2RX7) led to increase in intracellular calcium level that regulates the extracellular release of ATP. ATP was able to regulate calcimycin-induced autophagy through P2RX7 in an autocrine fashion. Blocking of either P2RX7 expression by 1-[N,O-bis(5-Isoquinolinesulfonyl)-N-methyl-L-tyrosyl]-4-phenylpiperazine (KN-62) or reducing intracellular calcium levels by 1,2-Bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid tetra (acetoxy-methyl) ester (BAPTA-AM) abrogated the antimycobacterial activity of calcimycin. Taken together, these results showed that calcimycin exerts its antimycobacterial effect by regulating intracellular calcium-dependent ATP release that induces autophagy in a P2RX7 dependent manner. CONCLUSION - Calcimycin exerts its antimycobacterial effect by regulating intracellular calcium-dependent ATP release that induces autophagy in a P2RX7 dependent manner. SIGNIFICANCE - Understanding the mechanism of autophagy by calcimycin may provide an attractive target for the control of mycobacterial infection, which will help in developing better therapeutic interventions against TB.