Citron kinase controls abscission through RhoA and Anillin.

The small GTPase RhoA plays a crucial role in the different stages of cytokinesis, including contractile ring formation, cleavage furrow ingression, and midbody abscission. Citron kinase (CIT-K), a protein required for cytokinesis and conserved from insects to mammals, is currently considered a cytokinesis-specific effector of active RhoA. In agreement with previous observations, we show here that, as in Drosophila cells, CIT-K is specifically required for abscission in mammalian cells. However, in contrast with the current view, we provide evidence that CIT-K is an upstream regulator rather than a downstream effector of RhoA during late cytokinesis. In addition, we show that CIT-K is capable of physically and functionally interacting with the actin-binding protein anillin. Active RhoA and anillin are displaced from the midbody in CIT-K-depleted cells, while only anillin, but not CIT-K, is affected if RhoA is inactivated in late cytokinesis. The overexpression of CIT-K and of anillin leads to abscission delay. However, the delay produced by CIT-K overexpression can be reversed by RhoA inactivation, while the delay produced by anillin overexpression is RhoA-independent. Altogether, these results indicate that CIT-K is a crucial abscission regulator that may promote midbody stability through active RhoA and anillin

miR-7b-3p exerts a dual role after spinal cord injury, by supporting plasticity and neuroprotection at cortical level

Spinal cord injury (SCI) affects 6 million people worldwide with no available treatment. Despite research advances, the inherent poor regeneration potential of the central nervous system remains a major hurdle. Small RNAs (sRNAs) 19–33 nucleotides in length are a set of non-coding RNA molecules that regulate gene expression and have emerged as key players in regulating cellular events occurring after SCI. Here we profiled a class of sRNA known as microRNAs (miRNAs) following SCI in the cortex where the cell bodies of corticospinal motor neurons are located. We identified miR-7b-3p as a candidate target given its significant upregulation after SCI in vivo and we screened by miRWalk PTM the genes predicted to be targets of miR-7b-3p (among which we identified Wipf2, a gene regulating neurite extension). Moreover, 16 genes, involved in neural regeneration and potential miR-7b-3p targets, were found to be downregulated in the cortex following SCI. We also analysed miR-7b-3p function during cortical neuron development in vitro: we observed that the overexpression of miR-7b-3p was important (1) to maintain neurons in a more immature and, likely, plastic neuronal developmental phase and (2) to contrast the apoptotic pathway; however, in normal conditions it did not affect the Wipf2 expression. On the contrary, the overexpression of miR-7b-3p upon in vitro oxidative stress condition (mimicking the SCI environment) significantly reduced the expression level of Wipf2, as observed in vivo, confirming it as a direct miR-7b-3p target. Overall, these data suggest a dual role of miR-7b-3p: (i) the induction of a more plastic neuronal condition/phase, possibly at the expense of the axon growth, (ii) the neuroprotective role exerted through the inhibition of the apoptotic cascade. Increasing the miR-7b-3p levels in case of SCI could reactivate in adult neurons silenced developmental programmes, supporting at the same time the survival of the axotomised neurons

The DCR protein TTC3 affects differentiation and Golgi compactness in neurons through specific actin-regulating pathways.

In neuronal cells, actin remodeling plays a well known role in neurite extension but is also deeply involved in the organization of intracellular structures, such as the Golgi apparatus. However, it is still not very clear which mechanisms may regulate actin dynamics at the different sites. In this report we show that high levels of the TTC3 protein, encoded by one of the genes of the Down Syndrome Critical Region (DCR), prevent neurite extension and disrupt Golgi compactness in differentiating primary neurons. These effects largely depend on the capability of TTC3 to promote actin polymerization through signaling pathways involving RhoA, ROCK, CIT-N and PIIa. However, the functional relationships between these molecules differ significantly if considering the TTC3 activity on neurite extension or on Golgi organization. Finally, our results reveal an unexpected stage-dependent requirement for F-actin in Golgi organization at different stages of neuronal differentiation

Citron Kinase Deficiency Leads to Chromosomal Instability and TP53-Sensitive Microcephaly

Mutations in citron (CIT), leading to loss or inactivation of the citron kinase protein (CITK), cause primary microcephaly in humans and rodents, associated with cytokinesis failure and apoptosis in neural progenitors. We show that CITK loss induces DNA damage accumulation and chromosomal instability in both mammals and Drosophila. CITK-deficient cells display "spontaneous" DNA damage, increased sensitivity to ionizing radiation, and defective recovery from radiation-induced DNA lesions. In CITK-deficient cells, DNA double-strand breaks increase independently of cytokinesis failure. Recruitment of RAD51 to DNA damage foci is compromised by CITK loss, and CITK physically interacts with RAD51, suggesting an involvement of CITK in homologous recombination. Consistent with this scenario, in doubly CitK and Trp53 mutant mice, neural progenitor cell death is dramatically reduced; moreover, clinical and neuroanatomical phenotypes are remarkably improved. Our results underscore a crucial role of CIT in the maintenance of genomic integrity during brain development

p140Cap regulates memory and synaptic plasticity through Src-mediated and citron-N-mediated actin reorganization.

Abstract A major challenge in the neuroscience field is the identification of molecules and pathways that control synaptic plasticity and memory. Dendritic spines play a pivotal role in these processes, as the major sites of excitatory synapses in neuronal communication. Previous studies have shown that the scaffold protein p140Cap localizes into dendritic spines and that its knockdown negatively modulates spine shape in culture. However, so far, there is no information on its in vivo relevance. By using a knock-outmousemodel, we here demonstrate that p140Cap is a key element for both learning and synaptic plasticity. Indeed, p140Cap(-/-) mice are impaired in object recognition test, as well as in LTP and in LTD measurements. The in vivo effects of p140Cap loss are presumably attenuated by noncell-autonomous events, since primary neurons obtained from p140Cap(-/-) mice show a strong reduction in number of mushroom spines and abnormal organization of synapse-associated F-actin. These phenotypes are most likely caused by a local reduction of the inhibitory control of RhoA and of cortactin toward the actin-depolymerizing factor cofilin. These events can be controlled by p140Cap through its capability to directly inhibit the activation of Src kinase and by its binding to the scaffold protein Citron-N. Altogether, our results provide new insight into how protein associated with dynamic microtubules may regulate spine actin organization through interaction with postsynaptic density components

Citron Kinase Deficiency Leads to Chromosomal Instability and TP53-Sensitive Microcephaly

Mutations in citron (CIT), leading to loss or inactivation of the citron kinase protein (CITK), cause primary microcephaly in humans and rodents, associated with cytokinesis failure and apoptosis in neural progenitors. We show that CITK loss induces DNA damage accumulation and chromosomal instability in both mammals and Drosophila. CITK-deficient cells display “spontaneous” DNA damage, increased sensitivity to ionizing radiation, and defective recovery from radiation-induced DNA lesions. In CITK-deficient cells, DNA double-strand breaks increase independently of cytokinesis failure. Recruitment of RAD51 to DNA damage foci is compromised by CITK loss, and CITK physically interacts with RAD51, suggesting an involvement of CITK in homologous recombination. Consistent with this scenario, in doubly CitK and Trp53 mutant mice, neural progenitor cell death is dramatically reduced; moreover, clinical and neuroanatomical phenotypes are remarkably improved. Our results underscore a crucial role of CIT in the maintenance of genomic integrity during brain development