Biology subjects

Kubo, Y.

Publications and source records attributed to Kubo, Y..

24 records · Page 2Linked to original sources

bioRxiv2020

Voltage-clamp fluorometry analysis of structural rearrangements of ATP-gated channel P2X2 upon hyperpolarization

The gating of the ATP-activated channel P2X2 has been shown to be dependent not only on [ATP] but also on membrane voltage, despite the absence of a canonical voltage-sensor domain. We aimed to investigate the structural rearrangements of the rat P2X2 during ATP- and voltage-dependent gating by voltage-clamp fluorometry technique. We observed fast and linearly voltage-dependent fluorescence intensity (F) changes at Ala337 and Ile341 in the TM2 domain, which could be due to the electrochromic effect, reflecting the presence of a converged electric field here. We also observed slow and voltage-dependent F changes at Ala337, which reflect the structural rearrangements. Furthermore, we identified that the interaction between Ala337 in TM2 and Phe44 in TM1, located in close proximity in the ATP-bound open state, is critical for activation. Taken together, we propose that the voltage dependence of the interaction in the converged electric field underlies the voltage-dependent gating.

biophysics↗

bioRxiv2020

Neurons learn by predicting future activity.

Understanding how the brain learns may lead to machines with human-like intellectual capacities. However, learning mechanisms in the brain are still not well understood. Here we demonstrate that the ability of a neuron to predict its future activity may provide an effective mechanism for learning in the brain. We show that comparing a neurons predicted activity with the actual activity provides a useful learning signal for modifying synaptic weights. Interestingly, this predictive learning rule can be derived from a metabolic principle, where neurons need to minimize their own synaptic activity (cost), while maximizing their impact on local blood supply by recruiting other neurons. This reveals an unexpected connection that learning in neural networks could result from simply maximizing the energy balance by each neuron. We show how this mathematically derived learning rule can provide a theoretical connection between diverse types of brain-inspired algorithms, such as: Hebbs rule, BCM theory, temporal difference learning and predictive coding. Thus, this may offer a step toward development of a general theory of neuronal learning. We validated this predictive learning rule in neural network simulations and in data recorded from awake animals. We found that in the sensory cortex it is indeed possible to predict a neurons activity [~]10-20ms into the future. Moreover, in response to stimuli, cortical neurons changed their firing rate to minimize surprise: i.e. the difference between actual and expected activity, as predicted by our model. Our results also suggest that spontaneous brain activity provides "training data" for neurons to learn to predict cortical dynamics. Thus, this work demonstrates that the ability of a neuron to predict its future inputs could be an important missing element to understand computation in the brain.

neuroscience↗

bioRxiv2020

Mechanism of hERG inhibition by gating-modifier toxin, APETx1, deduced by functional characterization

Human ether-a-go-go-related gene potassium channel 1 (hERG) is a voltage-gated potassium channel, the voltage-sensing domain (VSD) of which is targeted by a gating-modifier toxin, APETx1. Although it is known that APETx1 inhibits hERG by stabilizing the resting state, it remains unclear where and how APETx1 interacts with the VSD in the resting state. Here, we prepared a recombinant APETx1, which is structurally and functionally equivalent to the natural product. Electrophysiological analyses using wild type and mutants of APETx1 and hERG revealed that their hydrophobic residues, in addition to a previously reported acidic hERG residue, play key roles in the inhibition of hERG by APETx1. Docking models of the APETx1-VSD complex that satisfy the results of mutational analysis suggest a molecular recognition mode between APETx1 and the resting state of hERG; this would provide a structural basis for designing ligands that control hERG function by binding to the VSD.

molecular biology↗

bioRxiv2020

Arabidopsis SMN2/HEN2, encoding DEAD-box RNA helicase, governs proper expression of the resistance gene SMN1/RPS6 and is involved in dwarf, autoimmune phenotypes of mekk1 and mpk4

In Arabidopsis thaliana, a mitogen-activated protein kinase pathway, MEKK1-MKK1/MKK2-MPK4, is important for basal resistance, and disruption of this pathway results in dwarf, autoimmune phenotypes. To elucidate the complex mechanisms activated by the disruption of this pathway, we have previously developed a mutant screening system based on a dwarf autoimmune line that overexpressed the N-terminal regulatory domain of MEKK1. Here, we report that the second group of mutants, smn2, had defects in the SMN2 gene, encoding a DEAD-box RNA helicase. SMN2 is identical to HEN2, whose function is vital for the nuclear RNA exosome because it provides non-ribosomal RNA specificity for RNA turnover, RNA quality control, and RNA processing. Aberrant SMN1/RPS6 transcripts were detected in smn2 and hen2 mutants. Disease resistance against Pseudomonas syringae pv. tomato DC3000 (hopA1), which is conferred by SMN1/RPS6, was decreased in smn2 mutants, suggesting a functional connection between SMN1/RPS6 and SMN2/HEN2. We produced double mutants mekk1smn2 and mpk4smn2 to determine whether the smn2 mutations suppress the dwarf, autoimmune phenotypes of the mekk1 and mpk4 mutants, as the smn1 mutations do. As expected, the mekk1 and mpk4 phenotypes were suppressed by the smn2 mutations. These results suggested that SMN2 is involved in proper function of SMN1/RPS6. The GO enrichment analysis using RNA-seq data showed that defense genes were downregulated in smn2, suggesting positive contribution of SMN2 to genome-wide expression of defense-related genes. In conclusion, this study provides novel insight into plant immunity via SMN2/HEN2, an essential component of the nuclear RNA exosome.

plant biology↗

bioRxiv2019

CAT1/SLC7A1 acts as a cellular receptor for bovine leukemia virus infection

Bovine leukemia virus (BLV) is the causative agent of enzootic bovine leukosis, the most common neoplastic disease of cattle, which is closely related to human T-cell leukemia viruses. BLV has spread worldwide and causes a serious problem for the cattle industry. The cellular receptor specifically binds with viral envelope glycoprotein (Env) and this attachment mediates cell fusion to lead virus entry. BLV Env reportedly binds to cationic amino acid transporter 1 (CAT1)/SLC7A1, but whether the CAT1/SLC7A1 is an actual receptor for BLV remains unknown. Here, we showed that CAT1 functioned as an infection receptor, interacting with BLV particles. Cells expressing undetectable CAT1 levels were resistant to BLV infection but became highly susceptible upon CAT1 overexpression. CAT1 exhibited specific binding to BLV particles on the cell surface and co-localized with the Env in endomembrane compartments and membrane. Knockdown of CAT1 in permissive cells significantly reduced binding to BLV particles and BLV infection. In addition, bovine serum with neutralizing activity from a BLV-infected cattle inhibited BLV particles. Expression of CAT1 from various species demonstrated no species-specificity for BLV infection, implicating CAT1 as a functional BLV receptor responsible for its broad host range. These findings provide insights for BLV infection and for developing new strategies for treating BLV and preventing its spread.\n\nAuthor SummaryBovine leukemia virus (BLV), which can infect a variety of animal species and induce lymphoma in cattle, is a member of the family Retroviridae. BLV induces huge economic losses by not only lymphoma but also subclinical forms of the disease. In addition, BLV is frequently used as an animal model of human T-cell leukemia virus (HTLV), as BLV has many similar characteristics to HTLV. Thus, understanding BLV pathogenesis contribute to resolve not only BLV-but also HTLV-induced problems. Retroviral envelope glycoprotein (Env) is specifically recognized by the cellular receptor at cell surface, which induces a conformational changes between viral and cell membrane to entry. Thus, the elucidation of cellular receptor for BLV infection is very important for virus entry. However, the BLV receptor has not been identified yet. In the current study, we found that BLV Env protein binds to cationic amino acid transporter 1 (CAT1)/SLC7A1 at cell surface, artificial expression of CAT1 in CAT1-negative cells confers the cells susceptible to BLV infection, and CAT1-silencing significantly reduces BLV infection, concluding that CAT1 is the BLV receptor. These findings will have far reaching great advantages of insights in the retrovirus study.

microbiology↗

bioRxiv2018

SLO potassium channels antagonize premature decision making in C. elegans

SummaryAnimals have to modify their behavior at the right timing to respond to changes in environments. Yet, the molecular and neural mechanisms regulating the timing of behavioral transition remain largely unknown. Performing forward genetics on a plasticity of thermotaxis behavior in C. elegans, we demonstrated that SLO potassium channels together with a cyclic nucleotide-gated channel CNG-3 determine the timing of the transition of temperature preference after shift of cultivation temperature. We further revealed that SLO and CNG-3 channels regulate the alteration in responsiveness of thermosensory neurons. Our results suggest that the regulation of sensory adaptation is a major determinant of the latency for animals to make decisions in changing behavior.\n\nHighlightsO_LISlo-1 and SLO-2 K+ channels decelerated transition of temperature preference in thermotaxis behavior after upshift of cultivation temperature\nC_LIO_LISLO K+ channels slowed down the adaptation of AFD thermosensory neuron to new cultivation temperature\nC_LIO_LIA cyclic nucleotide-gated channel CNG-3 functioned together with SLO-2\nC_LIO_LIThermotaxis serves as could be a model system for early onset epilepsies\nC_LI

neuroscience↗