The Role of LRRK2 in Regulating a Calcium-Dependent Pathway Involving Lysosomal Stores and NAADP

Objective/Rationale:
Our preliminary evidence indicates that LRRK2 may regulate acidic calcium stores, with downstream effects on autophagy as well as a variety of other calcium dependent cellular events. Thus, mutant LRRK2 may lead to aberrant calcium homeostasis, which could contribute to Parkinson´s disease (PD). Our primary objective is to gain a direct understanding of the link between endogenous LRRK2 and calcium stores in a human cell culture model.
Project Description:
We have found that transient overexpression of active LRRK2 leads to the calcium-dependent activation of a signal transduction cascade to increase autophagosome numbers, a reduction in the number of acidic lysosomes and an increased sensitivity of cells to stressors associated with abnormal protein degradation. These effects can be mimicked by the lysosomal calcium-mobilizing messenger NAADP, and can be reverted by an NAADP receptor antagonist or expression of dominant-negative receptor constructs. We will investigate whether endogenous LRRK2 regulates acidic calcium stores in human fibroblasts from control and mutant LRRK2 patients. We will also determine whether the NAADP receptor antagonist can revert possible LRRK2-mediated effects on calcium stores in mutant fibroblasts. In parallel, we will perform experiments to study a possible LRRK2-NAADP receptor interaction.
Relevance to Diagnosis/Treatment of Parkinson’s Disease:
It is possible that LRRK2 regulates lysosomal calcium homeostasis, which may comprise a major event contributing to the disease. Establishing a clear link between LRRK2 and NAADP-sensitive calcium channels on acidic stores may pave the way for alternative therapeutic strategies targeting downstream events of LRRK2, distinct from a classical kinase inhibitor approach.
Anticipated Outcome:
We expect to learn whether LRRK2 regulates lysosomal calcium stores in a human cell culture model. Such deregulation of calcium homeostasis may explain many of the currently known cellular consequences due to abnormal LRRK2 function. However, our studies have been performed using overexpression in cell lines, and it is critical to validate whether endogenous LRRK2 regulates cellular calcium stores in human cell culture models and whether this may be likely relevant to PD.