Sam50 Regulates PINK1-Parkin-Mediated Mitophagy by Controlling PINK1 Stability and Mitochondrial Morphology

PINK1 and Parkin mediate mitophagy, the cellular process that clears dysfunctional mitochondria. Mitophagy is regulated by mitochondrial dynamics, but the molecules linking these two processes remain poorly understood. Here, we show that Sam50, the core component of the sorting and assembly machinery (SAM), is a critical regulator of mitochondrial dynamics and PINK1-Parkin-mediated mitophagy. In response to Sam50 depletion, normal tubular mitochondria are first fragmented and subsequently merged into large spheres. Sam50 interacts with PINK1 to facilitate its processing and degradation. Depletion of Sam50 results in PINK1 accumulation, Parkin recruitment, and mitophagy. Interestingly, Sam50 deficiency induces a piecemeal mode of mitophagy that eliminates mitochondria “bit by bit” but spares mtDNA. In C. elegans, the Sam50 homolog gop-3 is required for the maintenance of mitochondrial morphology and mass. Our findings reveal that Sam50 directly links mitochondrial dynamics and mitophagy and that Sam50 depletion induces elimination of mitochondria without affecting mtDNA content

LAMOST observations in the Kepler field. Analysis of the stellar parameters measured with the LASP based on the low-resolution spectra

All of the 14 subfields of the Kepler field have been observed at least once with the Large Sky Area Multi-Object Fiber Spectroscopic Telescope (LAMOST, Xinglong Observatory, China) during the 2012-2014 observation seasons. There are 88,628 reduced spectra with SNR$_g$ (signal-to-noise ratio in g band) $\geq$ 6 after the first round (2012-2014) of observations for the LAMOST-Kepler project (LK-project). By adopting the upgraded version of the LAMOST Stellar Parameter pipeline (LASP), we have determined the atmospheric parameters ($T_{\rm eff}$ , $\log g$, and $\rm [Fe/H]$) and heliocentric radial velocity $v_{\rm rad}$ for 51,406 stars with 61,226 spectra. Compared with atmospheric parameters derived from both high-resolution spectroscopy and asteroseismology method for common stars in Huber et al. (2014), an external calibration of LASP atmospheric parameters was made, leading to the determination of external errors for the giants and dwarfs, respectively. Multiple spectroscopic observations for the same objects of the LK-project were used to estimate the internal uncertainties of the atmospheric parameters as a function of SNR$_g$ with the unbiased estimation method. The LASP atmospheric parameters were calibrated based on both the external and internal uncertainties for the giants and dwarfs, respectively. A general statistical analysis of the stellar parameters leads to discovery of 106 candidate metal-poor stars, 9 candidate very metal-poor stars, and 18 candidate high-velocity stars. Fitting formulae were obtained segmentally for both the calibrated atmospheric parameters of the LK-project and the KIC parameters with the common stars. The calibrated atmospheric parameters and radial velocities of the LK-project will be useful for studying stars in the Kepler field.Comment: 53 pages, 21 figures, 5 tables, Accepted for publication by ApJ

LAMOST Observations in the Kepler Field. II. Database of the Low-resolution Spectra from the Five-year Regular Survey

The LAMOST-Kepler (LK-) project was initiated to use the Large Sky Area Multi-Object Fiber Spectroscopic Telescope (LAMOST) to make spectroscopic follow-up observations for the targets in the field of the Kepler mission. The Kepler field is divided into 14 subfields that are adapted to the LAMOST circular field with a diameter of 5°. During the regular survey phase of LAMOST, the LK-project took data from 2012 June to 2017 June and covered all 14 subfields at least twice. In particular, we describe in this paper the second Data Release of the LK-project, including all spectra acquired through 2015 May-2017 June together with the first round observations of the LK-project from 2012 June to 2014 September. The LK-project now counts 227,870 spectra of 156,390 stars, among which we have derived atmospheric parameters ({log}g, T eff, and [Fe/H]) and heliocentric radial velocity for 173,971 spectra of 126,172 stars. These parameters were obtained with the most recent version of the LAMOST Stellar Parameter Pipeline v 2.9.7. Nearly one half, namely 76,283 targets, are observed both by the LAMOST and Kepler telescopes. These spectra, establishing a large spectroscopy library, will be useful for the entire astronomical community, particularly for planetary science and stellar variability on Kepler targets. Based on observations collected with the Large Sky Area Multi-Object Fiber spectroscopic Telescope (LAMOST), which is located at the Xinglong Observatory, China

Regulation of endocytic recycling in Caenorhabditis elegans

Eukaryotic endocytic pathway is important for the uptake, sorting, and the subsequential recycling or degradation processes of various cargos. It has been shown that the RME-1/EHD1 is a critical regulator of endocytic recycling. In C. elegans and MDCK cells, small GTPase RAB-10 has been specifically implicated in clathrin-independent cargo recycling. Alternatively, some cargos will be recycled from sorting endosomes to Golgi, and retromer complex was shown to be crucial for this transport in yeast and mammalian cells. In our studies, we analyzed RME-1 and RAB-10 regulated recycling pathway, exploring additional players in the process. First, we demonstrated that ALX-1 is required for endocytic recycling of specific basolateral cargo. The interaction of ALX-1 with RME-1 is required for this recycling process. In our yeast two-hybrid screen for RAB-10-interacting proteins, EHBP-1 and Arf6 GAP/CNT-1 were recovered and revealed to function together with RAB-10 regulating clathrin-independent cargo recycling. Loss of either EHBP-1 or CNT-1 produced rab-10-like cargo transport defects. Furthermore, we showed that EHBP-1 functions, as an unconventional effector, upstream of RAB-10. Nevertheless, similar to canonical Rab effectors, CNT-1 requires RAB-10 for the proper endosome localization. Collectively, our results demonstrate the functional connections of ALX-1/RME-1 and RAB-10/EHBP-1/CNT-1, provided insights into the detail mechanisms of RME-1 and RAB-10 recycling regulation. We also studied the retromer regulated retrograde transport in C. elegans. We demonstrated the physical interaction of RME-8 with retromer component ALX-1. Additionally, we showed that loss-of-function in rme-8 or snx-1, or depletion of C. elegans Hsc70 (HSP-1) by RNAi, disrupts endosome to Golgi transport of the retromer-dependent cargo protein MIG-14. Furthermore, we identified a previously unsuspected mechanism for the regulation of endosomal clathrin that is required for retrograde transport. We showed that loss of either RME-8, SNX-1 or HSP-1 leads to endosomal clathrin dynamics defect. Our work indicates that, through RME-8 and Hsc70, the retromer acts to limit clathrin accumulation, a prerequisite for the recycling of retrograde cargo.Ph.D.Includes abstractVitaIncludes bibliographical referencesby Anbing Sh

Melatonin Mitigates Kainic Acid-Induced Neuronal Tau Hyperphosphorylation and Memory Deficits through Alleviating ER Stress

Kainic acid (KA) exposure causes neuronal degeneration featured by Alzheimer-like tau hyperphosphorylation and memory deficits. Melatonin (Mel) is known to protect hippocampal neurons against KA-induced damage. However, the underlying mechanisms remain elusive. In the current study, we investigated the protective effect of melatonin on KA-induced tau hyperphosphorylation by focusing on endoplasmic reticulum (ER) stress-mediated signaling pathways. By using primary hippocampal neurons and mouse brain, we showed that KA treatment specifically induced ER stress and activated GSK-3β and CDK5, two major kinases responsible for tau phosphorylation. Inhibition of ER stress efficiently inactivated GSK-3β and CDK5. Mechanistically, we found that KA-induced ER stress significantly activated calpain, a calcium-dependent protease. Inhibition of ER stress or calpain leads to the reduction in KA-induced GSK-3β and CDK5 activities and tau phosphorylation. Moreover, GSK-3β or CDK5 inhibition failed to downregulate ER stress efficiently, suggesting that ER stress functions upstream of GSK-3β or CDK5. Notably, our results revealed that melatonin acts against KA-induced neuronal degeneration and tau hyperphosphorylation via easing ER stress, further highlighting the protective role of melatonin in the KA-induced neuronal defects

A Novel Requirement for C. elegans Alix/ALX-1 in RME-1-Mediated Membrane Transport

SummaryBackgroundAlix/Bro1p family proteins have recently been identified as important components of multivesicular endosomes (MVEs) and are involved in the sorting of endocytosed integral membrane proteins, interacting with components of the ESCRT complex, the unconventional phospholipid LBPA, and other known endocytosis regulators. During infection, Alix can be co-opted by enveloped retroviruses, including HIV, providing an important function during virus budding from the plasma membrane. In addition, Alix is associated with the actin cytoskeleton and might regulate cytoskeletal dynamics.ResultsHere we demonstrate a novel physical interaction between the only apparent Alix/Bro1p family protein in C. elegans, ALX-1, and a key regulator of receptor recycling from endosomes to the plasma membrane, called RME-1. The analysis of alx-1 mutants indicates that ALX-1 is required for the endocytic recycling of specific basolateral cargo in the C. elegans intestine, a pathway previously defined by the analysis of rme-1 mutants. The expression of truncated human Alix in HeLa cells disrupts the recycling of major histocompatibility complex class I, a known Ehd1/RME-1-dependent transport step, suggesting the phylogenetic conservation of this function. We show that the interaction of ALX-1 with RME-1 in C. elegans, mediated by RME-1/YPSL and ALX-1/NPF motifs, is required for this recycling process. In the C. elegans intestine, ALX-1 localizes to both recycling endosomes and MVEs, but the ALX-1/RME-1 interaction appears to be dispensable for ALX-1 function in MVEs and/or late endosomes.ConclusionsThis work provides the first demonstration of a requirement for an Alix/Bro1p family member in the endocytic recycling pathway in association with the recycling regulator RME-1

Major contribution of the 3/6/7 class of TRPC channels to myocardial ischemia/reperfusion and cellular hypoxia/reoxygenation injuries

The injury phase after myocardial infarcts occurs during reperfusion and is a consequence of calcium release from internal stores combined with calcium entry, leading to cell death by apoptopic and necrotic processes. The mechanism(s) by which calcium enters cells has(ve) not been identified. Here, we identify canonical transient receptor potential channels (TRPC) 3 and 6 as the cation channels through which most of the damaging calcium enters cells to trigger their death, and we describe mechanisms activated during the injury phase. Working in vitro with H9c2 cardiomyoblasts subjected to 9-h hypoxia followed by 6-h reoxygenation (H/R), and analyzing changes occurring in areas-at-risk (AARs) of murine hearts subjected to a 30-min ischemia followed by 24-h reperfusion (I/R) protocol, we found: (i) that blocking TRPC with SKF96365 significantly ameliorated damage induced by H/R, including development of the mitochondrial permeability transition and proapoptotic changes in Bcl2/BAX ratios; and (ii) that AAR tissues had increased TUNEL+ cells, augmented Bcl2/BAX ratios, and increased p(S240)NFATc3, p(S473)AKT, p(S9)GSK3β, and TRPC3 and -6 proteins, consistent with activation of a positive-feedback loop in which calcium entering through TRPCs activates calcineurin-mediated NFATc3-directed transcription of TRPC genes, leading to more Ca2+ entry. All these changes were markedly reduced in mice lacking TRPC3, -6, and -7. The changes caused by I/R in AAR tissues were matched by those seen after H/R in cardiomyoblasts in all aspects except for p-AKT and p-GSK3β, which were decreased after H/R in cardiomyoblasts instead of increased. TRPC should be promising targets for pharmacologic intervention after cardiac infarcts.Fil: He, Xiju. Huazhong University Of Science And Technology; ChinaFil: Li, Shoutian. Huazhong University Of Science And Technology; ChinaFil: Liu, Benju. Huazhong University Of Science And Technology; ChinaFil: Susperreguy, Sebastian. Pontificia Universidad Católica Argentina ; ArgentinaFil: Formoso, Karina. Pontificia Universidad Católica Argentina ; ArgentinaFil: Yao, Jinghong. Huazhong University Of Science And Technology; ChinaFil: Kang, Jinsong. Huazhong University Of Science And Technology; ChinaFil: Shi, Anbing. Huazhong University Of Science And Technology; ChinaFil: Birnbaumer, Lutz. Pontificia Universidad Católica Argentina "Santa María de los Buenos Aires". Instituto de Investigaciones Biomédicas. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas; ArgentinaFil: Liao, Yanhong. Huazhong University Of Science And Technology; Chin

An EHBP-1-SID-3-DYN-1 axis promotes membranous tubule fission during endocytic recycling.

The ACK family tyrosine kinase SID-3 is involved in the endocytic uptake of double-stranded RNA. Here we identified SID-3 as a previously unappreciated recycling regulator in the Caenorhabditis elegans intestine. The RAB-10 effector EHBP-1 is required for the endosomal localization of SID-3. Accordingly, animals with loss of SID-3 phenocopied the recycling defects observed in ehbp-1 and rab-10 single mutants. Moreover, we detected sequential protein interactions between EHBP-1, SID-3, NCK-1, and DYN-1. In the absence of SID-3, DYN-1 failed to localize at tubular recycling endosomes, and membrane tubules breaking away from endosomes were mostly absent, suggesting that SID-3 acts synergistically with the downstream DYN-1 to promote endosomal tubule fission. In agreement with these observations, overexpression of DYN-1 significantly increased recycling transport in SID-3-deficient cells. Finally, we noticed that loss of RAB-10 or EHBP-1 compromised feeding RNAi efficiency in multiple tissues, implicating basolateral recycling in the transport of RNA silencing signals. Taken together, our study demonstrated that in C. elegans intestinal epithelia, SID-3 acts downstream of EHBP-1 to direct fission of recycling endosomal tubules in concert with NCK-1 and DYN-1