Cerebral palsy (CP), a neurodevelopmental disorder characterized by irreversible, nonprogressive central motor dysfunction, is commonly associated with prematurity or perinatal brain injury. However, accumulating evidence suggests deleterious genomic variants may contribute to CP in addition to environmental insults. To identify genes contributing to risk for CP, we performed whole-exome sequencing on 250 parent-offspring CP trios. We identified a significant contribution of damaging de novo mutations (DNMs), especially in genes that are intolerant to loss of function mutations. Eight genes had multiple, independently-arising damaging DNMs, including two novel CP-associated genes, FBXO31 and RHOB, and four genes previously implicated in cerebral palsy phenotypes, TUBA1A, CTNNB1, SPAST, and ATL1. Functional experiments, including molecular and biochemical assays and patient fibroblast studies indicate that the recurrent RHOB mutation identified in patients enhances Rho effector binding in the active state and that the FBXO31 mutation leads to elevated levels of cyclin D. Analysis of candidate CP risk genes highlighted genetic overlap with hereditary spastic paraplegia as well as intellectual disability, autism, and epilepsy, converging with epidemiologic findings. Computational network analysis of risk genes identified significant enrichment of Rho GTPase, extracellular matrix, focal adhesions, cytoskeleton, and cell projection pathways. CP risk genes in Rho GTPase, cytoskeleton and cell projection pathways were found to play an important role in neuromotor development via a Drosophila reverse genetics screen. Based on enrichment analysis, we estimate that an excess of damaging de novo and inherited recessive variants collectively account for ~14% of the cases in our cohort, whereas perinatal asphyxia is currently estimated to occur in 8-10% of CP cases. Together, these findings provide evidence for the role of genetically-mediated dysregulation of early brain connectivity in CP.

Circadian timekeeping is a ubiquitous mechanism that enables organisms to maintain temporal coordination between internal biological processes and time of the local environment. The molecular basis of circadian rhythms lies in a set of transcription-translation feedback loops (TTFLs) that drives the rhythmic transcription of core clock genes, whose level and phase of expression serve as the marker of circadian time. However, it has become increasingly evident that additional regulatory mechanisms impinge upon the TTFLs to govern the properties and behavior of the circadian clock. Such mechanisms include changes in chromatin architecture, interactions with other transcription factor networks, post-transcriptional control by RNA modifications, alternative splicing and microRNAs, and post-translational regulation of subcellular trafficking and protein degradation. In this review, we will summarize the current knowledge of circadian clock regulation-from transcriptional to post-translational-drawing from literature pertaining to the Drosophila and murine circadian systems.

PIDD1 encodes p53-Induced Death Domain protein 1, which acts as a sensor surveilling centrosome numbers and p53 activity in mammalian cells. Early results also suggest a role in DNA damage response where PIDD1 may act as a cell-fate switch, through interaction with RIP1 and NEMO/IKKg, activating NF-κB signaling for survival, or as an apoptosis-inducing protein by activating caspase-2. Biallelic truncating mutations in CRADD—the protein bridging PIDD1 and caspase-2—have been reported in intellectual disability (ID), and in a form of lissencephaly. Here, we identified five families with ID from Iran, Pakistan, and India, with four different biallelic mutations in PIDD1, all disrupting the Death Domain (DD), through which PIDD1 interacts with CRADD or RIP1. Nonsense mutations Gln863* and Arg637* directly disrupt the DD, as does a missense mutation, Arg815Trp. A homozygous splice mutation in the fifth family is predicted to disrupt splicing upstream of the DD, as confirmed using an exon trap. In HEK293 cells, we show that both Gln863* and Arg815Trp mutants fail to co-localize with CRADD, leading to its aggregation and mis-localization, and fail to co-precipitate CRADD. Using genome-edited cell lines, we show that these three PIDD1 mutations all cause loss of PIDDosome function. Pidd1 null mice show decreased anxiety, but no motor abnormalities. Together this indicates that PIDD1 mutations in humans may cause ID (and possibly lissencephaly) either through gain of function or secondarily, due to altered scaffolding properties, while complete loss of PIDD1, as modeled in mice, may be well tolerated or is compensated for.

Circadian locomotor output cycles kaput (CLOCK) is a nuclear transcription factor that is a component of the central autoregulatory feedback loop that governs the generation of biological rhythms. Homozygous Clock mutant mice contain a truncated CLOCK(Δ19) protein within somatic cells, subsequently causing an impaired ability to rhythmically transactivate circadian genes. The present study sought to investigate whether the Clock mutation affects mitochondrial physiology within skeletal muscle, as well as the responsiveness of these mutant animals to adapt to a chronic voluntary endurance training protocol. Within muscle, Clock mutant mice displayed 44% and 45% reductions in peroxisome proliferator-activated receptor-γ coactivator 1-α (PGC-1α) and mitochondrial transcription factor-A protein content, respectively, and an accompanying 16% decrease in mitochondrial content, as determined by cytochrome c oxidase enzyme activity. These decrements contributed to a 50% decrease in exercise tolerance in Clock mutant mice. Interestingly, the Clock mutation did not appear to alter subsarcolemmal or intermyofibrillar mitochondrial respiration within muscle or systemic glucose tolerance. Daily locomotor activity levels were similar between wild-type and Clock mutant mice throughout the training protocol. Endurance training ameliorated the decrease in PGC-1α protein expression and mitochondrial content in the Clock mutant mice, eliciting a 2.9-fold improvement in exercise tolerance. Thus our data suggest that a functional CLOCK protein is essential to ensure the maintenance of mitochondrial content within muscle although the absence of a functional CLOCK protein does not impair the ability of animals to adapt to chronic exercise.