There are mutations in the kinase domain and in the ROC-COR bi-domain that may affect either kinase or GTPase activity. The most prevalent LRRK2 mutation, G2019S, lies within the Mg +2 -binding motif of the kinase domain, and has been shown to increase the kinase activity of LRKK2, both in heterologous and autophosphorylation assays. However, whether other mutations in LRRK2 affect kinase activity is controversial. A caveat about several published studies is that autophosphorylation was used to measure kinase activity. Although such assays can be helpful in vitro, it is unclear whether autophosphorylation is Cortodoxone physiologically relevant. An alternative to autophosphorylation as an assay for kinase activity is to use generic substrates such as myelin basic protein, which generally give similar results to autophosphorylation. Several heterologous LRRK2 substrates have been suggested for LRRK2 including moesin, b-tubulin and MAPKKK substrates MKK3/6 or MKK4/7. Whether any of these substrates are physiologically relevant is currently unclear. Recently Imai et al. reported 4E-BP as a potential substrate of LRRK2. 4E-BP is an interactor of the eukaryotic protein translation initiation factor eIF4E, which in turn binds to capped mRNA species, promoting their translation. Binding of 4E-BP to eIF4E prevents the latter being active and, therefore, 4E-BP is a repressor of protein translation. Oxidative stress and other Cefpiramide sodium stimuli that impact protein translation affect phosphorylation of 4E-BP. Imai et al. proposed that LRRK2 modulates this system by phosphorylating 4E-BP at a specific site, which then acts as a stimulus for further phosphorylation by other kinases at secondary sites including S65/S70. There was a modest decrease in phosphorylation of 4E-BP T37/T46 and S65 when LRRK2 levels were knocked down with RNAi. Overexpression of 4E-BP rescued the effects of LRRK mutants in vivo using Drosophila models. Similarly, Tain et al have shown that phospho-4E-BP levels are decreased in a homozygous knockout model of drosophila LRRK. Collectively these data are supportive of 4E-BP being a substrate for LRRK2 or its Drosophila homologue, dLRRK.