Leucine-rich repeat kinase 2 positively regulates inflammation and down-regulates NF-κB p50 signaling in cultured microglia cells

Abstract

BackgroundOver-activated microglia and chronic neuroinflammation contribute to dopaminergic neuron degeneration and progression of Parkinson’s disease (PD). Leucine-rich repeat kinase 2 (LRRK2), a kinase mutated in autosomal dominantly inherited and sporadic PD cases, is highly expressed in immune cells, in which it regulates inflammation through a yet unclear mechanism.MethodsHere, using pharmacological inhibition and cultured Lrrk2−/− primary microglia cells, we validated LRRK2 as a positive modulator of inflammation and we investigated its specific function in microglia cells.ResultsInhibition or genetic deletion of LRRK2 causes reduction of interleukin-1β and cyclooxygenase-2 expression upon lipopolysaccharide-mediated inflammation. LRRK2 also takes part of the signaling trigged by α-synuclein fibrils, which culminates in induction of inflammatory mediators. At the molecular level, loss of LRRK2 or inhibition of its kinase activity results in increased phosphorylation of nuclear factor kappa-B (NF-κB) inhibitory subunit p50 at S337, a protein kinase A (PKA)-specific phosphorylation site, with consequent accumulation of p50 in the nucleus.ConclusionsTaken together, these findings point to a role of LRRK2 in microglia activation and sustainment of neuroinflammation and in controlling of NF-κB p50 inhibitory signaling. Understanding the molecular pathways coordinated by LRRK2 in activated microglia cells after pathological stimuli such us fibrillar α-synuclein holds the potential to provide novel targets for PD therapeutics.

Mutations in LRRK2 impair NF-κB pathway in iPSC-derived neurons

“…Discrepancies may be partly due to the use of different cellular systems and stimulation conditions and, in the case of LRRK2 inhibitors, to off-target effects. On the other hand, knockdown experiments of endogenous LRRK2 expression in primary microglia and immortalized immune cell lines down-regulated inflammatory signaling, even in the absence of a pro-inflammatory stimulus [14, 15, 44, 45], which is also in agreement with our findings in LRRK2-silenced fibroblasts [6]. In neurons, we found a differential regulation of NF-κB transcriptional targets, which may be dependent on cell-specific factors.…”

Leucine-Rich Repeat Kinase 2 Influences Fate Decision of Human Monocytes Differentiated from Induced Pluripotent Stem Cells

“…By contrast, LRRK2 is associated with activation of immune cells [14,54] and increased inflammatory responses have been found in peripheral immune cells of LRRK2 mutant mice by us and others [13,15,18]. Several studies linked LRRK2-deficiency or LRRK2 inhibition to decreased cytokine release from activated mouse macrophages [55] and microglia [15,20,56,57], whereas Liu et al [58] reported increased cytokine release from activated bone marrow-derived macrophages isolated from LRRK2-deficient mice. These studies, however, differed regarding the cell populations and applied stimuli under investigation, indicating that LRRK2 possibly exerts different effects under distinct experimental conditions.…”

Cross-talk between LRRK2 and PKA: implication for Parkinson's disease?

“…Supporting our results, Kim et al [38] showed that BV2 cells with LRRK2 knockdown display reduced NF-κB transcription activity and increased p50:p50 homodimer bound to NF-κB target sites compared with wild-type cells, further supporting the notion that LRRK2 controls NF-κB p50 inhibitory signaling. Different groups found a reduced inflammation in microglia cells with LRRK2 knockdown or pharmacological inhibition after inflammatory stimuli [14,[38][39][40]. In this regard, we propose that the enhanced phosphorylation of NF-κB p50 inhibitory subunit bound to DNA might be one molecular mechanism underlying the reduction in NF-κB target genes transcription upon inflammatory stimuli.…”

“…Recently, we demonstrated that primary mouse microglia with Lrrk2 genetic deletion and immortalized BV2 microglia upon LRRK2 pharmacological inhibition exhibit increased PKA-mediated phosphorylation of nuclear NF-κB p50 subunit [14]. NF-κB p50:p50 homodimer phosphorylated by PKA is constitutively imported into the nucleus [36], possesses a high affinity for DNA [21] and is essential to repress the transcription of NF-κB target genes in the absence of extracellular stimuli [37].…”

“…LRRK2 has been linked to distinct signaling cascades, including MAPK [7], Rac/PAK [8,9], Wnt [10], Akt [11], TLR (reviewed in ref. [12]) and cAMP-PKA ( protein kinase A) [13,14] pathways. All these pathways can affect multiple cellular processes, but those in which LRRK2 has been convincingly associated are autophagy, cytoskeletal dynamics, membrane trafficking, mitochondria and inflammation (reviewed in ref.…”