LRRK2 secretion in exosomes is regulated by 14-3-3

Abstract

Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene cause late-onset Parkinson's disease (PD). Emerging evidence suggests a role for LRRK2 in the endocytic pathway. Here, we show that LRRK2 is released in extracellular microvesicles (i.e. exosomes) from cells that natively express LRRK2. LRRK2 localizes to collecting duct epithelial cells in the kidney that actively secrete exosomes into urine. Purified urinary exosomes contain LRRK2 protein that is both dimerized and phosphorylated. We provide a quantitative proteomic profile of 1673 proteins in urinary exosomes and find that known LRRK2 interactors including 14-3-3 are some of the most abundant exosome proteins. Disruption of the 14-3-3 LRRK2 interaction with a 14-3-3 inhibitor or through acute LRRK2 kinase inhibition potently blocks LRRK2 release in exosomes, but familial mutations in LRRK2 had no effect on secretion. LRRK2 levels were overall comparable but highly variable in urinary exosomes derived from PD cases and age-matched controls, although very high LRRK2 levels were detected in some PD affected cases. We further characterized LRRK2 exosome release in neurons and macrophages in culture, and found that LRRK2-positive exosomes circulate in cerebral spinal fluid (CSF). Together, these results define a pathway for LRRK2 extracellular release, clarify one function of the LRRK2 14-3-3 interaction and provide a foundation for utilization of LRRK2 as a biomarker in clinical trials.

Transcellular spreading of huntingtin aggregates in theDrosophilabrain

“…Although there is abundant evidence that spreading occurs through synaptic connections, other potential mechanisms include spreading between cells via exosomes (43,44) or tunneling nanotubes (45). In the current study, we find unique patterns of spreading when mutant Htt is expressed in different subsets of neurons in the brain.…”

Parkinson-related LRRK2 mutation R1441C/G/H impairs PKA phosphorylation of LRRK2 and disrupts its interaction with 14-3-3

“…binding. Furthermore, recent data from Fraser et al (22) suggest a regulatory function of 14-3-3 binding in controlling extracellular release of LRRK2. Dysregulation of 14-3-3/client protein interaction has been shown to facilitate the development of several human disorders (23,24), and an influence of the pathogenic mutation R1441G on LRRK2/14-3-3 interaction has been demonstrated (12,19).…”

Proteomic profile of maternal-aged blastocoel fluid suggests a novel role for ubiquitin system in blastocyst quality

“…neXtProt.org, www.exocarta.org, microvesicle.org, geneontology.org, www.proteinatlas. org), and dataset [29]. Gene products previously detected at the transcription level in blastocystis or embyonic tissue are listed Tables 1, 2, 3, and 4 and S2-according to microarray datasets and gene expression databases (http://genomewidepdb.proteomix.org or https://www.ncbi.nlm.nih.gov/geoprofiles) [16,30].…”

“…All consenting patients undergoing fertility treatment at the Humanitas Fertility Center, Humanitas Research Hospital, Rozzano, Italy, with supernumerary blastocysts were included in the study irrespective of the woman's age (range years [26][27][28][29][30][31][32][33][34][35][36][37][38][39][40][41], infertility diagnosis (female or male), ovarian stimulation protocol (use of GnRH agonist or antagonist protocols in combination with recombinant and/or highly purified urinary gonadotropins), or performance of standard IVF or intracytoplasmic sperm injection (ICSI). The study was part of a trial on blastocoel fluid collection in order to test the safety of this procedure on pregnancy outcome [22].…”