Dynamic and redundant regulation of LRRK2 and LRRK1 expression

Institute for Cell Engineering and Department of Neurology, Johns Hopkins University School of Medicine, Baltimore, USA.
BMC Neuroscience (Impact Factor: 2.67). 02/2007; 8(1):102. DOI: 10.1186/1471-2202-8-102
Source: PubMed


Mutations within the leucine-rich repeat kinase 2 (LRRK2) gene account for a significant proportion of autosomal-dominant and some late-onset sporadic Parkinson's disease. Elucidation of LRRK2 protein function in health and disease provides an opportunity for deciphering molecular pathways important in neurodegeneration. In mammals, LRRK1 and LRRK2 protein comprise a unique family encoding a GTPase domain that controls intrinsic kinase activity. The expression profiles of the murine LRRK proteins have not been fully described and insufficiently characterized antibodies have produced conflicting results in the literature.
Herein, we comprehensively evaluate twenty-one commercially available antibodies to the LRRK2 protein using mouse LRRK2 and human LRRK2 expression vectors, wild-type and LRRK2-null mouse brain lysates and human brain lysates. Eleven antibodies detect over-expressed human LRRK2 while four antibodies detect endogenous human LRRK2. In contrast, two antibodies recognize over-expressed mouse LRRK2 and one antibody detected endogenous mouse LRRK2. LRRK2 protein resides in both soluble and detergent soluble protein fractions. LRRK2 and the related LRRK1 genes encode low levels of expressed mRNA species corresponding to low levels of protein both during development and in adulthood with largely redundant expression profiles.
Despite previously published results, commercially available antibodies generally fail to recognize endogenous mouse LRRK2 protein; however, several antibodies retain the ability to detect over-expressed mouse LRRK2 protein. Over half of the commercially available antibodies tested detect over-expressed human LRRK2 protein and some have sufficient specificity to detect endogenous LRRK2 in human brain. The mammalian LRRK proteins are developmentally regulated in several tissues and coordinated expression suggest possible redundancy in the function between LRRK1 and LRRK2.

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    • "However, others have found that neurite phenotypes are robust only during the first week in vitro (Sepulveda et al., 2013). Comparatively little LRRK2 is expressed during this period, whereas LRRK2 levels ∼double between the first and second week, both in vitro and in vivo (Biskup et al., 2007; Piccoli et al., 2011), during which time glutamatergic "
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    ABSTRACT: Mutations in Leucine-Rich Repeat Kinase-2 (LRRK2) result in familial Parkinson's disease and the G2019S mutation alone accounts for up to 30% in some ethnicities. Despite this, the function of LRRK2 is largely undetermined although evidence suggests roles in phosphorylation, protein interactions, autophagy and endocytosis. Emerging reports link loss of LRRK2 to altered synaptic transmission, but the effects of the G2019S mutation upon synaptic release in mammalian neurons are unknown. To assess wild type and mutant LRRK2 in established neuronal networks, we conducted immunocytochemical, electrophysiological and biochemical characterization of >3 week old cortical cultures of LRRK2 knock-out, wild-type overexpressing and G2019S knock-in mice. Synaptic release and synapse numbers were grossly normal in LRRK2 knock-out cells, but discretely reduced glutamatergic activity and reduced synaptic protein levels were observed. Conversely, synapse density was modestly but significantly increased in wild-type LRRK2 overexpressing cultures although event frequency was not. In knock-in cultures, glutamate release was markedly elevated, in the absence of any change to synapse density, indicating that physiological levels of G2019S LRRK2 elevate probability of release. Several pre-synaptic regulatory proteins shown by others to interact with LRRK2 were expressed at normal levels in knock-in cultures; however, synapsin 1 phosphorylation was significantly reduced. Thus, perturbations to the pre-synaptic release machinery and elevated synaptic transmission are early neuronal effects of LRRK2 G2019S. Furthermore, the comparison of knock-in and overexpressing cultures suggests that one copy of the G2019S mutation has a more pronounced effect than an ~3-fold increase in LRRK2 protein. Mutant-induced increases in transmission may convey additional stressors to neuronal physiology that may eventually contribute to the pathogenesis of Parkinson's disease.
    Frontiers in Cellular Neuroscience 09/2014; 8:301. DOI:10.3389/fncel.2014.00301 · 4.29 Impact Factor
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    • "Both LRRK1 and LRRK2 mRNA have been reported to be widely expressed in adult human brain, with comparative studies indicating relatively strong expression of both in cortical regions and the hippocampus while at least in one study, the striatum demonstrated relatively high levels of LRRK2 but not LRRK1 (Westerlund et al. 2008; Dachsel et al. 2010). Also, other studies have demonstrated significantly higher levels of LRRK2 in the brain relative to LRRK1 (Biskup et al. 2007). Interestingly, LRRK1 has been proposed to regulate endocytosis of epidermal growth factor receptor (EGFR) via formation of a LRRK1-Grb2-EGFR complex (Hanafusa et al. 2011), while LRRK2 did not appear to form such a complex in this study, suggesting that the two have functionally diverged. "
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    ABSTRACT: Mutations in the catalytic Roc-COR and kinase domains of leucine-rich repeat kinase 2 (LRRK2) are a common cause of familial Parkinson's disease (PD). LRRK2 mutations cause PD with age-related penetrance and clinical features identical to late-onset sporadic PD. Biochemical studies support an increase in LRRK2 kinase activity and a decrease in GTPase activity for kinase domain and Roc-COR mutations, respectively. Strong evidence exists that LRRK2 toxicity is kinase-dependent leading to extensive efforts to identify selective and brain-permeable LRRK2 kinase inhibitors for clinical development. Cell and animal models of PD indicate that LRRK2 mutations affect vesicular trafficking, autophagy, protein synthesis and cytoskeletal function. Although some of these biological functions are affected consistently by most disease-linked mutations, others are not and it is currently unclear how mutations that produce variable effects on LRRK2 biochemistry and function all commonly result in the degeneration and death of dopamine neurons. LRRK2 is typically present in Lewy bodies and its toxicity in mammalian models appears to be dependent on the presence of α-synuclein, which is elevated in human iPS-derived dopamine neurons from patients harboring LRRK2 mutations. Here, we summarize biochemical and functional studies of LRRK2 and its mutations and focus on aberrant vesicular trafficking and protein synthesis as two leading mechanisms underlying LRRK2-linked disease.This article is protected by copyright. All rights reserved.
    Journal of Neurochemistry 09/2014; 131(5). DOI:10.1111/jnc.12949 · 4.28 Impact Factor
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    • "Some studies have demonstrated localization of LRRK2 to α-synuclein pathology (Miklossy et al., 2006; Alegre-Abarrategui et al., 2008; Qing et al., 2009; Sharma et al., 2011) whilst others have not (Giasson et al., 2006; Higashi et al., 2007; Melrose et al., 2007; Waxman et al., 2009). Studies comparing different LRRK2 antibodies have shown that discrepancies in LRRK2 tissue localization likely occurs through use of antibodies unsuitable for immunohistochemistry (Biskup et al., 2007; Melrose et al., 2007; Davies et al., 2013). Indeed, recent data using more rigorous methods shows LRRK2 and α-synuclein co-localize in a small proportion of PD pathologies (Guerreiro et al., 2013). "
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    ABSTRACT: Substantial evidence implicates abnormal protein kinase function in various aspects of Parkinson's disease (PD) etiology. Elevated phosphorylation of the PD-defining pathological protein, α-synuclein, correlates with its aggregation and toxic accumulation in neurons, whilst genetic missense mutations in the kinases PTEN-induced putative kinase 1 and leucine-rich repeat kinase 2, increase susceptibility to PD. Experimental evidence also links kinases of the phosphoinositide 3-kinase and mitogen-activated protein kinase signaling pathways, amongst others, to PD. Understanding how the levels or activities of these enzymes or their substrates change in brain tissue in relation to pathological states can provide insight into disease pathogenesis. Moreover, understanding when and where kinase dysfunction occurs is important as modulation of some of these signaling pathways can potentially lead to PD therapeutics. This review will summarize what is currently known in regard to the expression of these PD-implicated kinases in pathological human postmortem brain tissue.
    Frontiers in Molecular Neuroscience 06/2014; 7:57. DOI:10.3389/fnmol.2014.00057 · 4.08 Impact Factor
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