Targeting specific kinase substrates rescues increased colitis severity induced by the Crohn's disease-linked LRRK2-N2081D variant.

LRRK2 contains a kinase domain where the N2081D Crohn's disease (CD) risk and the G2019S Parkinson's disease (PD) pathogenic variants are located. It is not clear how the N2081D variant increases CD risk or how these adjacent mutations give rise to distinct disorders. To investigate the pathophysiology of the CD-linked LRRK2 N2081D variant, we generated a knock-in (KI) mouse model and compared its effects with those of the LRRK2-G2019S mutation. Lrrk2N2081D KI mice demonstrated heightened sensitivity to induced colitis, resulting in more severe intestinal damage than in Lrrk2G2019S KI and WT mice. Analysis of colon tissue revealed distinct mutation-dependent LRRK2 RAB substrate phosphorylation, with significantly elevated phosphorylated RAB10 levels in Lrrk2N2081D mice. In cells, we demonstrated that the N2081D mutation activates LRRK2 through a mechanism distinct from that of LRRK2-G2019S. We also found that proinflammatory stimulation enhances LRRK2 kinase activity, leading to mutation-dependent differences in RAB phosphorylation and inflammatory responses in dendritic cells (DCs). Finally, we show that knockout of Rab12, but not pharmacological LRRK2 kinase inhibition, significantly reduced colitis severity in Lrrk2N2081D mice. Our study characterizes the pathogenic mechanisms of LRRK2-linked CD, highlights structural and functional differences between disease-associated LRRK2 variants, and suggests RAB proteins as promising therapeutic targets for modulating LRRK2 activity in CD treatment.