Классификация экзонов генов человека с использованием алгоритмов автоматического выбора атрибутов экзонов

Volkov AV, Yatskov NN, Grinev VV Classification of exons of human genes using algorithms for automatic selection of exon attributesСекция 2. МЕТОДЫ И ТЕХНОЛОГИИ МАТЕМАТИЧЕСКОГО И ИМИТАЦИОННОГО МОДЕЛИРОВАНИЯ СИСТЕ

An intellectual-disability-associated mutation of the transcriptional regulator NACC1 impairs glutamatergic neurotransmission

Advances in genome sequencing technologies have favored the identification of rare de novo mutations linked to neurological disorders in humans. Recently, a de novo autosomal dominant mutation in NACC1 was identified (NM_052876.3: c.892C > T, NP_443108.1; p.Arg298Trp), associated with severe neurological symptoms including intellectual disability, microcephaly, and epilepsy. As NACC1 had never before been associated with neurological diseases, we investigated how this mutation might lead to altered brain function. We examined neurotransmission in autaptic glutamatergic mouse neurons expressing the murine homolog of the human mutant NACC1, i.e., Nacc1-R284W. We observed that expression of Nacc1-R284W impaired glutamatergic neurotransmission in a cell-autonomous manner, likely through a dominant negative mechanism. Furthermore, by screening for Nacc1 interaction targets in the brain, we identified SynGAP1, GluK2A, and several SUMO E3 ligases as novel Nacc1 interaction partners. At a biochemical level, Nacc1-R284W exhibited reduced binding to SynGAP1 and GluK2A, and also showed greatly increased SUMOylation. Ablating the SUMOylation of Nacc1-R284W partially restored its interaction with SynGAP1 but did not restore binding to GluK2A. Overall, these data indicate a role for Nacc1 in regulating glutamatergic neurotransmission, which is substantially impaired by the expression of a disease-associated Nacc1 mutant. This study provides the first functional insights into potential deficits in neuronal function in patients expressing the de novo mutant NACC1 protein

Two distinct secretory vesicle–priming steps in adrenal chromaffin cells

The calcium-dependent activator proteins for secretion, CAPS1 and CAPS2, facilitate syntaxin opening during synaptic vesicle priming

Molecular Dissection of Neurobeachin Function at Excitatory Synapses

Spines are small protrusions from dendrites where most excitatory synapses reside. Changes in number, shape, and size of dendritic spines often reflect changes of neural activity in entire circuits or at individual synapses, making spines key structures of synaptic plasticity. Neurobeachin is a multidomain protein with roles in spine formation, postsynaptic neurotransmitter receptor targeting and actin distribution. However, the contributions of individual domains of Neurobeachin to these functions is poorly understood. Here, we used mostly live cell imaging and patch-clamp electrophysiology to monitor morphology and function of spinous synapses in primary hippocampal neurons. We demonstrate that a recombinant full-length Neurobeachin from humans can restore mushroom spine density and excitatory postsynaptic currents in neurons of Neurobeachin-deficient mice. We then probed the role of individual domains of Neurobeachin by comparing them to the full-length molecule in rescue experiments of knockout neurons. We show that the combined PH-BEACH domain complex is highly localized in spine heads, and that it is sufficient to restore normal spine density and surface targeting of postsynaptic AMPA receptors. In addition, we report that the Armadillo domain facilitates the formation of filopodia, long dendritic protrusions which often precede the development of mature spines, whereas the PKA-binding site appears as a negative regulator of filopodial extension. Thus, our results indicate that individual domains of Neurobeachin sustain important and specific roles in the regulation of spinous synapses. Since heterozygous mutations in Neurobeachin occur in autistic patients, the results will also improve our understanding of pathomechanism in neuropsychiatric disorders associated with impairments of spine function

Secretory Vesicle Priming by CAPS Is Independent of Its SNARE-Binding MUN Domain

Priming of secretory vesicles is a prerequisite for their Ca2+-dependent fusion with the plasma membrane. The key vesicle priming proteins, Munc13s and CAPSs, are thought to mediate vesicle priming by regulating the conformation of the t-SNARE syntaxin, thereby facilitating SNARE complex assembly. Munc13s execute their priming function through their MUN domain. Given that the MUN domain of Ca2+-dependent activator protein for secretion (CAPS) also binds syntaxin, it was assumed that CAPSs prime vesicles through the same mechanism as Munc13s. We studied naturally occurring splice variants of CAPS2 in CAPS1/CAPS2-deficient cells and found that CAPS2 primes vesicles independently of its MUN domain. Instead, the pleckstrin homology domain of CAPS2 seemingly is essential for its priming function. Our findings indicate a priming mode for secretory vesicles. This process apparently requires membrane phospholipids, does not involve the binding or direct conformational regulation of syntaxin by MUN domains of CAPSs, and is therefore not redundant with Munc13 action

The Morphological and Molecular Nature of Synaptic Vesicle Priming at Presynaptic Active Zones

SummarySynaptic vesicle docking, priming, and fusion at active zones are orchestrated by a complex molecular machinery. We employed hippocampal organotypic slice cultures from mice lacking key presynaptic proteins, cryofixation, and three-dimensional electron tomography to study the mechanism of synaptic vesicle docking in the same experimental setting, with high precision, and in a near-native state. We dissected previously indistinguishable, sequential steps in synaptic vesicle active zone recruitment (tethering) and membrane attachment (docking) and found that vesicle docking requires Munc13/CAPS family priming proteins and all three neuronal SNAREs, but not Synaptotagmin-1 or Complexins. Our data indicate that membrane-attached vesicles comprise the readily releasable pool of fusion-competent vesicles and that synaptic vesicle docking, priming, and trans-SNARE complex assembly are the respective morphological, functional, and molecular manifestations of the same process, which operates downstream of vesicle tethering by active zone components