Expression of microRNAs in cerebrospinal fluid of dogs with central nervous system disease

Abstract In this pilot study we investigated the expression of 14 microRNAs in the cerebrospinal fluid (CSF) of dogs with neoplastic, inflammatory and degenerative disorders affecting the central nervous system (CNS). CSF microRNA (miRNA) expression profiles were compared to those from dogs with neurological signs but no evidence of structural or inflammatory CNS disease. Seven miRNAs were easily detected in all samples: miR-10b-5p, miR-19b, miR-21-5p, miR-30b-5p, miR-103a-3p, miR-124, and miR-128-3p. Expression of miR-10b-5p was significantly higher in the neoplastic group compared to other groups. There was no relation between miRNA expression and either CSF nucleated cell count or CSF protein content. Higher expression of miR-10b-5p in the neoplastic group is consistent with previous reports in human medicine where aberrant expression of miR-10b is associated with various neoplastic diseases of the CNS

CRYPTOCHROMES confer robustness, not rhythmicity, to circadian timekeeping

Circadian rhythms are a pervasive property of mammalian cells, tissues and behaviour, ensuring physiological adaptation to solar time. Models of cellular timekeeping revolve around transcriptional feedback repression, whereby CLOCK and BMAL1 activate the expression of PERIOD (PER) and CRYPTOCHROME (CRY), which in turn repress CLOCK/BMAL1 activity. CRY proteins are therefore considered essential components of the cellular clock mechanism, supported by behavioural arrhythmicity of CRY‐deficient (CKO) mice under constant conditions. Challenging this interpretation, we find locomotor rhythms in adult CKO mice under specific environmental conditions and circadian rhythms in cellular PER2 levels when CRY is absent. CRY‐less oscillations are variable in their expression and have shorter periods than wild‐type controls. Importantly, we find classic circadian hallmarks such as temperature compensation and period determination by CK1δ/ε activity to be maintained. In the absence of CRY‐mediated feedback repression and rhythmic Per2 transcription, PER2 protein rhythms are sustained for several cycles, accompanied by circadian variation in protein stability. We suggest that, whereas circadian transcriptional feedback imparts robustness and functionality onto biological clocks, the core timekeeping mechanism is post‐translational

Macromolecular condensation buffers intracellular water potential

Optimum protein function and biochemical activity critically depends on water availability because solvent thermodynamics drive protein folding and macromolecular interactions1. Reciprocally, macromolecules restrict the movement of 'structured' water molecules within their hydration layers, reducing the available 'free' bulk solvent and therefore the total thermodynamic potential energy of water, or water potential. Here, within concentrated macromolecular solutions such as the cytosol, we found that modest changes in temperature greatly affect the water potential, and are counteracted by opposing changes in osmotic strength. This duality of temperature and osmotic strength enables simple manipulations of solvent thermodynamics to prevent cell death after extreme cold or heat shock. Physiologically, cells must sustain their activity against fluctuating temperature, pressure and osmotic strength, which impact water availability within seconds. Yet, established mechanisms of water homeostasis act over much slower timescales2,3; we therefore postulated the existence of a rapid compensatory response. We find that this function is performed by water potential-driven changes in macromolecular assembly, particularly biomolecular condensation of intrinsically disordered proteins. The formation and dissolution of biomolecular condensates liberates and captures free water, respectively, quickly counteracting thermal or osmotic perturbations of water potential, which is consequently robustly buffered in the cytoplasm. Our results indicate that biomolecular condensation constitutes an intrinsic biophysical feedback response that rapidly compensates for intracellular osmotic and thermal fluctuations. We suggest that preserving water availability within the concentrated cytosol is an overlooked evolutionary driver of protein (dis)order and function