Expression Analysis of Lrrk1, Lrrk2 and Lrrk2 Splice Variants in Mice

Abstract

Missense mutations in the leucine-rich repeat kinase 2 gene ( LRRK2 ) are linked to autosomal dominant forms of Parkinson’s disease (PD). In order to get insights into the physiological role of Lrrk2 , we examined the distribution of Lrrk2 mRNA and different splice variants in the developing murine embryo and the adult brain of Mus musculus. To analyse if the Lrrk2 -paralog, Lrrk1, may have redundant functions in PD-development, we also compared Lrrk1 and Lrrk2 expression in the same tissues. Using radioactive in situ hybridization, we found ubiquitous expression of both genes at low level from embryonic stage E9.5 onward, which progressively increased up until birth. The developing central nervous system (CNS) displayed no prominent Lrrk2 mRNA signals at these time-points. However, in the entire postnatal brain Lrrk2 became detectable, showing strongest level in the striatum and the cortex of adult mice; Lrrk1 was only detectable in the mitral cell layer of the olfactory bulb. Thus, due to the non-overlapping expression patterns, a redundant function of Lrrk2 and Lrrk1 in the pathogenesis of PD seems to be unlikely. Quantification of Lrrk2 mRNA and protein level in several brain regions by real-time PCR and Western blot verified the striatum and cortex as hotspots of postnatal Lrrk2 expression. Strong expression of Lrrk2 is mainly found in neurons, specifically in the dopamine receptor 1 (DRD1a) and 2 (DRD2)-positive subpopulations of the striatal medium spiny neurons. Finally, we identified 2 new splice-variants of Lrrk2 in RNA-samples from various adult brain regions and organs: a variant with a skipped exon 5 and a truncated variant terminating in an alternative exon 42a. In order to identify the origin of these two splice variants, we also analysed primary neural cultures independently and found cell-specific expression patterns for these variants in microglia and astrocytes.