The Role of BTBD9 in Striatum and Restless Legs Syndrome

Abstract

Restless legs syndrome (RLS) is a sensory-motor neurological disorder characterized by uncomfortable sensations in the extremities, generally at night, which is often relieved by movements. Genome-wide association studies (GWAS) have identified mutations in BTBD9 conferring a higher risk of RLS. Knockout of the BTBD9 homolog in mice (Btbd9) and fly results in motor restlessness and sleep disruption. Clinical studies have found RLS patients have structural and functional abnormalities in the striatum; however, whether and how striatal pathology contributes to the pathogenesis of RLS is not known. Here, we used fMRI to map regions of altered synaptic activity in basal ganglia of systematic Btbd9 knock-out (KO) mice. We further dissected striatal circuits using patch-clamp electrophysiological recordings in brain slices. Two different mouse models were generated to test the effect of specific knockout of Btbd9 in either striatal medium spiny neurons (MSNs) or cholinergic interneurons (ChIs) using the electrophysiological recording, motor and sensory behavioral tests. We found that Btbd9 KO mice showed enhanced neural activity in the striatum, increased postsynaptic currents in the MSNs, and decreased excitability of the striatal ChIs. Knocking out Btbd9 specifically in the striatal MSNs, but not the ChIs, led to rest-phase specific motor restlessness, sleep disturbance, and increased thermal sensation in mice, which are consistent with results obtained from the Btbd9 KO mice. Our data establish the role of Btbd9 in regulating the activity of striatal neurons. Increased activity of the striatal MSNs, possibly through modulation by the striatal ChIs, contributes to the pathogenesis of RLS.

The CRL3BTBD9 E3 ubiquitin ligase complex targets TNFAIP1 for degradation to suppress cancer cell migration

“…BTBD9, a BTB domain-containing protein, is significantly associated with susceptibility to sleep disturbance and restless leg syndrome (RLS). [37][38][39][40] However, there has been no study about its role in cancer or its role as an adaptor of CRL3 E3 ligases that modulates the degradation of tumor suppressors or oncoproteins. In this study, BTBD9 was first identified as an adaptor of CRL3 that regulates the ubiquitination and degradation of its substrate protein TNFAIP1, as demonstrated by the following: (1) TNFAIP1 and BTBD9 interacted, (2) downregulation of BTBD9 delayed the turnover of TNFAIP1, and (3) BTBD9 knockdown diminished the polyubiquitination of TNFAIP1.…”

Deficiency of Meis1, a transcriptional regulator, in mice and worms: Neurochemical and behavioral characterizations with implications in the restless legs syndrome

“…Six Meis1 KO males and five WT males with an average age of 9 months were tested in the continuous open field experiment as previously described (Lyu, DeAndrade, et al, 2019, 2020; Lyu, Doroodchi, Xing, et al, 2020; Lyu, Xing, et al, 2019; Lyu, Xing, DeAndrade, Perez, Yokoi, et al, 2020; Lyu, Xing, DeAndrade, Perez, Zhang, et al, 2020). The sample size was determined according to (Lyu, Xing, et al, 2019). The average weight of the animals is 34.5 g. Briefly, mice were maintained on a 12 LD cycle for 7 days in the VersaMax Legacy Open Field activity box (Omnitech Electronics, year of purchase, 2012).…”

“…Recordings of dopamine neurons were performed with three Meis1 KO males and three WT males with an average age of 10 months (Lyu, Doroodchi, Xing, et al, 2020). The sample size was determined according to (Lyu, Xing, et al, 2019 and 10 glucose, pH = 7.3 with KOH, osmolality = 290-300 mOsm).…”

Probing the relationship between BTBD9 and MEIS1 in C. elegans and mouse

“…Heterozygous Btbd9 KO (Lyu et al, 2019) were bred with Meis1 KO animals (Meneely et al, 2018) to generate double heterozygotes, which were bred with heterozygous Btbd9 KO mice to generate experimental mice. Behavioral tests were conducted as described (Lyu et al, 2019) and analyzed by SAS GENMOD or mixed model ANOVA. Western blot and quantitative RT-PCR were performed and analyzed as described (Yokoi et al, 2015) using striatal tissues (supplementary material).…”

“…What does "4 repeats for each genotype" mean? Does this mean the last 4 days that were used in the analysis according to Lyu et al 2019 or did the mice spend four times seven days in the running wheel?…”