Cyclic di-AMP in bacterial homeostasis: from membrane-bound synthesis and stress sensing to transport

Abstract

The thesis investigates the synthesis and transport of cyclic di-AMP in Lactococcus lactis.Chapter1 introduces cyclic di-AMP's role in bacterial osmotic homeostasis. Chapter2 focuses on the function and structure of the synthetase for cyclic di-AMP, CdaA, revealing that the membrane-bound, tetrameric form exhibits significantly higher activity than its soluble catalytic domain, a finding supported by cryo-EM structural analysis. Chapter3 explores why CdaA is membrane-associated, finding its activity is subtly influenced by complex membrane biophysical properties like curvature and surface charge, rather than a single factor.Chapter4 investigates cyclic di-AMP transport, studying putative exporters Lmo2588 and Llmg1210-1211. Initial antiport/symport hypotheses were inconclusive on the basis of experiments in proteoliposomes, but pH-dependent uptake was observed in crude membrane vesicles. The reason for the proteoliposome system's failure remains unclear. These transporters also appear to confer cholic acid resistance, though passive diffusion could not be ruled out.In conclusion, this work provides structural insights into membrane-bound CdaA's high activity and reveals complex membrane-dependent regulation. While the identity of the cylic di-AMP transporter remains unresolved, the findings challenge previous genetic-based claims