Multiple Autism-Linked Genes Mediate Synapse Elimination via Proteasomal Degradation of a Synaptic Scaffold PSD-95

SummaryThe activity-dependent transcription factor myocyte enhancer factor 2 (MEF2) induces excitatory synapse elimination in mouse neurons, which requires fragile X mental retardation protein (FMRP), an RNA-binding protein implicated in human cognitive dysfunction and autism. We report here that protocadherin 10 (Pcdh10), an autism-spectrum disorders gene, is necessary for this process. MEF2 and FMRP cooperatively regulate the expression of Pcdh10. Upon MEF2 activation, PSD-95 is ubiquitinated by the ubiquitin E3 ligase murine double minute 2 (Mdm2) and then binds to Pcdh10, which links it to the proteasome for degradation. Blockade of the Pcdh10-proteasome interaction inhibits MEF2-induced PSD-95 degradation and synapse elimination. In FMRP-lacking neurons, elevated protein levels of eukaryotic translation elongation factor 1 α (EF1α), an Mdm2-interacting protein and FMRP target mRNA, sequester Mdm2 and prevent MEF2-induced PSD-95 ubiquitination and synapse elimination. Together, our findings reveal roles for multiple autism-linked genes in activity-dependent synapse elimination

Homeostatic regulation of extracellular signal-regulated kinase 1/2 activity and axonal Kv7.3 expression by prolonged blockade of hippocampal neuronal activity

Homeostatic plasticity encompasses the mechanisms by which neurons stabilize their synaptic strength and excitability in response to prolonged and destabilizing changes in their network activity. Prolonged activity blockade leads to homeostatic scaling of action potential (AP) firing rate in hippocampal neurons in part by decreased activity of N-Methyl-D-Aspartate receptors and subsequent transcriptional down-regulation of potassium channel genes including KCNQ3 which encodes Kv7.3. Neuronal Kv7 channels are mostly heterotetramers of Kv7.2 and Kv7.3 subunits and are highly enriched at the axon initial segment (AIS) where their current potently inhibits repetitive and burst firing of APs. However, whether a decrease in Kv7.3 expression occurs at the AIS during homeostatic scaling of intrinsic excitability and what signaling pathway reduces KCNQ3 transcript upon prolonged activity blockade remain unknown. Here, we report that prolonged activity blockade in cultured hippocampal neurons reduces the activity of extracellular signal-regulated kinase 1/2 (ERK1/2) followed by a decrease in the activation of brain-derived neurotrophic factor (BDNF) receptor, Tropomyosin receptor kinase B (TrkB). Furthermore, both prolonged activity blockade and prolonged pharmacological inhibition of ERK1/2 decrease KCNQ3 and BDNF transcripts as well as the density of Kv7.3 and ankyrin-G at the AIS. Collectively, our findings suggest that a reduction in the ERK1/2 activity and subsequent transcriptional down-regulation may serve as a potential signaling pathway that links prolonged activity blockade to homeostatic control of BDNF-TrkB signaling and Kv7.3 density at the AIS during homeostatic scaling of AP firing rate

PKCε Stimulated Arginine Methylation of RIP140 for Its Nuclear-Cytoplasmic Export in Adipocyte Differentiation

Receptor interacting protein 140 (RIP140) is a versatile transcriptional co-repressor that plays roles in diverse metabolic processes including fat accumulation in adipocytes. Previously we identified three methylated arginine residues in RIP140, which rendered its export to the cytoplasm; but it was unclear what triggered RIP140 arginine methylation.In this study, we determined the activated PKCepsilon as the specific trigger for RIP140 arginine methylation and its subsequent export. We identified two PKCepsilon-phosphorylated residues of RIP140, Ser-102 and Ser-1003, which synergistically stimulated direct binding of RIP140 by 14-3-3 that recruited protein arginine methyl transferase 1 to methylate RIP140. The methylated RIP140 then preferentially recruited exportin 1 for nuclear export. As a result, the nuclear gene-repressive activity of RIP140 was reduced. In RIP140 null adipocyte cultures, the defect in fat accumulation was effectively rescued by the phosphorylation-deficient mutant RIP140 that resided predominantly in the nucleus, but less so by the phospho-mimetic RIP140 that was exported to the cytoplasm.This study uncovers a novel means, via a cascade of protein modifications, to inactivate, or suppress, the nuclear action of an important transcription coregulator RIP140, and delineates the first specific phosphorylation-arginine methylation cascade that could alter protein subcellular distribution and biological activity

Novel roles of ER stress in repressing neural activity and seizures through Mdm2- and p53-dependent protein translation.

Seizures can induce endoplasmic reticulum (ER) stress, and sustained ER stress contributes to neuronal death after epileptic seizures. Despite the recent debate on whether inhibiting ER stress can reduce neuronal death after seizures, whether and how ER stress impacts neural activity and seizures remain unclear. In this study, we discovered that the acute ER stress response functions to repress neural activity through a protein translation-dependent mechanism. We found that inducing ER stress promotes the expression and distribution of murine double minute-2 (Mdm2) in the nucleus, leading to ubiquitination and down-regulation of the tumor suppressor p53. Reduction of p53 subsequently maintains protein translation, before the onset of translational repression seen during the latter phase of the ER stress response. Disruption of Mdm2 in an Mdm2 conditional knockdown (cKD) mouse model impairs ER stress-induced p53 down-regulation, protein translation, and reduction of neural activity and seizure severity. Importantly, these defects in Mdm2 cKD mice were restored by both pharmacological and genetic inhibition of p53 to mimic the inactivation of p53 seen during ER stress. Altogether, our study uncovered a novel mechanism by which neurons respond to acute ER stress. Further, this mechanism plays a beneficial role in reducing neural activity and seizure severity. These findings caution against inhibition of ER stress as a neuroprotective strategy for seizures, epilepsies, and other pathological conditions associated with excessive neural activity

Time variation of high-risk groups for liver function deteriorations within fluctuating long-term liver function after hepatic radiotherapy in patients with hepatocellular carcinoma

Abstract Purpose The purpose of this study is to find essential risk factors associated with liver function (LF) deteriorations within fluctuating long-term LF and their time-varying effects in patients with hepatocellular carcinoma (HCC) receiving hepatic radiotherapy and to identify high-risk groups for adverse LF deteriorations and their changes over time in facilitating the prevention of hepatic decompensation and the improvement of survival. Materials and methods A total of 133 HCC patients treated by hepatic radiotherapy were enrolled. A study design was conducted to convert posttreatment long-term LF with fluctuating levels over time to recurrent LF events using defined upgrades in a grading scale. The hazard ratios (HR) of pretreatment biochemical, demographic, clinical, and dosimetric factors in developing posttreatment LF events were estimated using the Cox model. Methodologies of the counting process approach, robust variance estimation, goodness-of-fit testing based on the Schoenfeld residuals, and time-dependent covariates in survival analysis were employed to handle the correlation within subjects and evaluate the time-varying effects during long-term follow-up. Results Baseline LF score before radiotherapy and gender were significant factors. Initial HR in developing LF events was 1.17 (95% CI 1.11–1.23; P < 0.001) for each increase of baseline LF score and kept almost constant over time (HR, 1.00; 95% CI 1.00–1.01; P = 0.065). However, no difference was observed regarding initial hazards for gender (HR, 1.00; 95% CI 0.64–1.56; P = 0.994), but the hazard for women got higher monthly over time compared with men (HR, 1.04; 95% CI 1.01–1.07; P = 0.006). Conclusions High-risk groups for adverse LF deteriorations after hepatic radiotherapy may change over time. Patients with poor baseline LF are vulnerable from the beginning. Women require prevention strategies and careful monitoring for deteriorations at a later stage

Tangent-based volumetric modulated arc therapy for advanced left breast cancer

Abstract Purpose To introduce the benefits of tangent-based volumetric modulated arc therapy (TVMAT), an innovative radiotherapy planning technique, compared with traditional volumetric modulated arc therapy (VMAT) for advanced left breast cancer needing nodal irradiation. Materials and methods Twenty-three patients with advanced left breast cancer who had received modified radical mastectomy (MRM) and needed adjuvant radiotherapy including nodal irradiation were assessed. Among 23 radiotherapy treatment plans, 17 plans were designed by using TVMAT technique and 6 plans were designed by using traditional VMAT. The main difference of TVMAT from VMAT was that the area of avoidance sector within specific degrees of angle that had no monitor unit (MU) delivery was used in the arc planning, including a total of 5 sectors in 5 partial arcs. The dosimetries of planning target volume (PTV), right breast, bilateral lungs, and heart between TVMAT and VMAT were compared. Results The conformity index (CI) and homogeneity index (HI) of PTV between two groups were statistically equivalent (CI: 0.98 ± 0.02 and 0.98 ± 0.03, P = 0.431; HI: 0.12 ± 0.03 and 0.11 ± 0.05, P = 0.177), which indicated that the treatment efficacy of the plans regarding TVMAT was compatible with VMAT. However, all neighboring organs at risk (OAR) showed a great percentage of reduction in mean doses (right breast: 53.1%, right lung: 37.7%, left lung: 8.8%, heart: 21.2%) and low dose parameters (V10: right breast: 72.3%, right lung: 86.1%, left lung: 12.5%, heart: 25.1%; V5: right breast: 56.5%, right lung: 28.3%, left lung: 12.7%, heart: 18.2%) by using TVMAT. Conclusion TVMAT greatly decreases the radiation doses delivered to the OAR with maintained therapeutic efficacy. It is highly recommended for treating breast cancer, especially for difficult cases with left side disease needing nodal irradiation

Quantitative analysis of respiration-induced motion of each liver segment with helical computed tomography and 4-dimensional computed tomography

Abstract Background To analyze the respiratory-induced motion of each liver segment using helical computed tomography (helical CT) and 4-dimensional computed tomography (4DCT), and to establish the individual segment expansion margin of internal target volume (ITV) to facilitate target delineation of tumors in different liver segments. Methods Twenty patients who received radiotherapy with CT-simulation scanning of the whole liver in both helical CT and 10-phase-gated 4DCT were investigated, including 2 patients with esophagus cancer, 4 with lung cancer, 10 with breast cancer, 2 with liver cancer, 1 with thymoma, and 1 with gastric diffuse large B-cell lymphoma (DLBCL). For each patient, 9 representative points were drawn on the helical CT images of liver segments 1, 2, 3, 4a, 4b, 5, 6, 7, and 8, respectively, and adaptively deformed to 2 phases of the 4DCT images at the end of inspiration (phase 0 CT) and expiration (phase 50 CT) in the treatment planning system. Using the amplitude of each point between phase 0 CT and phase 50 CT, we established quantitative data for the respiration-induced motion of each liver segment in 3-dimensional directions. Moreover, using the amplitude between the original helical CT and both 4DCT images, we rendered the individual segment expansion margin of ITV for hepatic target delineation to cover more than 95% of each tumor. Results The average amplitude (mean ± standard deviation) was 0.6 ± 3.0 mm in the left-right (LR) direction, 2.3 ± 2.4 mm in the anterior-posterior (AP) direction, and 5.7 ± 3.4 mm in the superior-inferior (SI) direction, respectively. All of the segments moved posteriorly and superiorly during expiration. Segment 7 had the largest amplitude in the SI direction, at 8.6 ± 3.4 mm. Otherwise, the segments over the lateral side, including segments 2, 3, 6, and 7, had greater excursion in the SI direction compared to the medial segments. To cover more than 95% of each tumor, the required expansion margin of ITV in the LR, AP, and SI directions were at least 2.5 mm, 2.5 mm, and 5.0 mm on average, respectively, with variations between different segments. Conclusions The greatest excursion occurred in liver segment 7, followed by the segments over the lateral side in the SI direction. The individual segment expansion margin of ITV is required to delineate targets for each segment and direction