Spliceosome component Usp39 contributes to hepatic lipid homeostasis through the regulation of autophagy.

Regulation of alternative splicing (AS) enables a single transcript to yield multiple isoforms that increase transcriptome and proteome diversity. Here, we report that spliceosome component Usp39 plays a role in the regulation of hepatocyte lipid homeostasis. We demonstrate that Usp39 expression is downregulated in hepatic tissues of non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH) subjects. Hepatocyte-specific Usp39 deletion in mice leads to increased lipid accumulation, spontaneous steatosis and impaired autophagy. Combined analysis of RNA immunoprecipitation (RIP-seq) and bulk RNA sequencing (RNA-seq) data reveals that Usp39 regulates AS of several autophagy-related genes. In particular, deletion of Usp39 results in alternative 5' splice site selection of exon 6 in Heat shock transcription factor 1 (Hsf1) and consequently its reduced expression. Importantly, overexpression of Hsf1 could attenuate lipid accumulation caused by Usp39 deficiency. Taken together, our findings indicate that Usp39-mediated AS is required for sustaining autophagy and lipid homeostasis in the liver.

Bud31-mediated alternative splicing is required for spermatogonial stem cell self-renewal and differentiation.

Alternative splicing (AS) is tightly regulated during cell differentiation and development. AS events are prevalent in the testis, but the splicing regulation in spermatogenesis remains unclear. Here we report that the spliceosome component Bud31 plays a crucial role during spermatogenesis in mice. Germ cell-specific knockout of Bud31 led to loss of spermatogonia and to male infertility. We further demonstrate that Bud31 is required for both spermatogonial stem cell pool maintenance and the initiation of spermatogenesis. SMART-seq revealed that deletion of Bud31 in germ cells causes widespread exon-skipping and intron retention. Particularly, we identified Cdk2 as one of the direct splicing targets of Bud31, knockout of Bud31 resulted in retention of the first intron of Cdk2, which led to a decrease in Cdk2 expression. Our findings suggest that Bud31-mediated AS within spermatogonial stem cells regulates the self-renewal and differentiation of male germ cells in mammals.

A multi-center, single-arm, phase II study of anlotinib plus paclitaxel and cisplatin as the first-line therapy of recurrent/advanced esophageal squamous cell carcinoma.

BACKGROUND: Anlotinib, a tyrosine kinase inhibitor, has shown encouraging anti-tumor activity in esophageal squamous cell carcinoma (ESCC). This study was designed to assess the efficacy and safety of anlotinib plus paclitaxel and cisplatin (TP) as first-line therapy for advanced ESCC. METHODS: In a multi-center, single-arm, phase II clinical trial, patients (aged > 18 years) with ESCC, which was judged to be locally advanced, recurrent, or metastatic, received 10 mg oral anlotinib once daily on days 1-14, 135 mg/m2 intravenous paclitaxel on day 1, and 60-75 mg/m2 intravenous cisplatin on days 1-3 every 3 weeks for a maximum of 4-6 cycles as the initial therapy in five centers in China. Subsequently, patients received anlotinib monotherapy (10 mg) as maintenance therapy until tumor progression or intolerable toxicity. The primary endpoint was progression-free survival (PFS). RESULTS: Forty-seven patients were enrolled in this study between October 2019 and March 2021. The median follow-up was 14.04 months (IQR, 9.30-19.38). Of 46 with assessable efficacy, the median PFS and median overall survival were 8.38 months (95% CI, 6.59-10.17) and 18.53 months (95% CI, 13.11-23.95), respectively. The objective response rate was 76.1% (95% CI, 61.2-87.4%), with 4 (8.7%) complete responses and 31 (67.4%) partial responses. The disease control rate was 91.3% (95% CI, 79.2-97.6%). The median duration of response was 6.80 months (95% CI, 4.52-9.08), and 1 patient had an ongoing response for 23 months. Subgroup analysis revealed no association between clinical factors and survival or response. Of the 47 patients with assessable safety, the main grade ≥ 3 treatment-emergent adverse events (TEAEs) were neutropenia (17.0%), bone marrow suppression (12.8%), and vomiting (10.6%). No treatment-related deaths or serious TEAEs were observed. Notably, higher c-Kit levels were an independent factor for superior PFS (HR = 0.032; 95% CI, 0.002-0.606; P = 0.022). CONCLUSIONS: The study demonstrated a manageable safety profile and durable clinical response of anlotinib plus TP as first-line therapy in advanced ESCC, which suggested a potential therapeutic option for this population. TRIAL REGISTRATION: ClinicalTrials.gov NCT04063683. Registered 21 August 2019.

Mixed conifer-broadleaf trees on arbuscular mycorrhizal and ectomycorrhizal communities in rhizosphere soil of different plantation stands in the temperate zone, Northeast China.

In comparison with ectomycorrhizal (EM) tree species, arbuscular mycorrhizal (AM) trees have different litter quality and nitrogen cycle modes, which may affect mycorrhizal colonization and the community composition and diversity. However, available studies addressing the mycorrhizal fungal colonization rate, diversity and community composition in mixed forest stands composed of AM and EM trees are rare. In the present study, we assessed litter quality, soil physicochemical properties and correlated them with mycorrhizal community characteristics in rhizosphere soils of monoculture and mixture plantation stands of AM tree species (Fraxinus mandschurica Rupr.) and EM tree species (Larix gmelinii Rupr., Picea koraiensis Nakai) in Northeast China. We hypothesized that (1) the effect of mixture pattern on mycorrhizal colonization rate and diversity would change with tree species, (2) the effect of mixture pattern on mycorrhizal community composition would be less pronounced in comparison with that of tree species. We found that mixture did not change AMF colonization rate regardless of mixture identity, whereas mixture and tree species exerted significant effects on EMF colonization rate. For AMF community, both M-AS (Fraxinus mandschurica Rupr. and Picea koraiensis Nakai) and M-AL (Fraxinus mandschurica Rupr. and Larix gmelinii Rupr.) mixtures significantly increased Pielou index and Simpson index, whereas only M-AS significantly increased Sobs. For EMF community, mixture significantly affected examined diversity indices except for Chao1. Mixture significantly shifted AMF and EMF community, and the magnitude was tree species dependent. The dominant genera in AMF and EMF communities in plantation stands were Glomus and Tomentella, respectively. The EnvFit analysis showed that the determinant factors of EMF community are soil moisture, pH, nitrate nitrogen content, dissolved organic nitrogen content, soil organic matter content, soil organic carbon/total nitrogen and litter carbon/total nitrogen. In conclusion, mixed conifer-broadleaf trees significantly changed soil physicochemical properties, litter quality as well as mycorrhizal fungi community diversity and composition.

Overexpression of HMGA1 confers radioresistance by transactivating RAD51 in cholangiocarcinoma.

Cholangiocarcinomas (CCAs) are rare but aggressive tumors of the bile ducts. CCAs are often diagnosed at an advanced stage and respond poorly to current conventional radiotherapy and chemotherapy. High mobility group A1 (HMGA1) is an architectural transcription factor that is overexpressed in multiple malignant tumors. In this study, we showed that the expression of HMGA1 is frequently elevated in CCAs and that the high expression of this gene is associated with a poor prognosis. Functionally, HMGA1 promotes CCA cell proliferation/invasion and xenograft tumor growth. Furthermore, HMGA1 transcriptionally activates RAD51 by binding to its promoter through two HMGA1 response elements. Notably, overexpression of HMGA1 promotes radioresistance whereas its knockdown causes radiosensitivity of CCA cells to X-ray irradiation. Moreover, rescue experiments reveal that inhibition of RAD51 reverses the effect of HMGA1 on radioresistance and proliferation/invasion. These findings suggest that HMGA1 functions as a novel regulator of RAD51 and confers radioresistance in cholangiocarcinoma.

[Root architecture and fine root characteristics of Juglans mandshurica saplings in different habitats in the secondary forest on the west slope of Zhangguangcailing, China]. / å¼ å¹¿æ‰å²­è¥¿å¡æ¬¡ç”Ÿæž—ä¸åŒç”Ÿå¢ƒèƒ¡æ¡ƒæ¥¸å¹¼æ ‘æ ¹ç³»æž„åž‹åŠç»†æ ¹ç‰¹å¾.

The whole root excavation method was used to examine root configuration of Juglans mandshurica, with the age of 5-6 years in three habitats (forest edge, gap, and canopy) in a secondary forest on the western part of Zhangguangcailing Mountains. Root structure and fine root function were measured. The root topological index, average joint length, cross-sectional area ratio before and after root branching were calculated and fine root chemical compositions were analyzed. Roots of J. mandshurica at forest edge tended to be dichotomous branch (Topological index:TI=0.68), that under the canopy were herringbone-like branch (TI=0.79), and the gap was between the two (TI=0.72). The average connection length of roots among the three habitats was not significant. The cross-sectional area ratio of roots before and after root branching in three habitats was 1.06, 1.04 and 1.07, respectively, which was not affected by root diameter, in accordance with the Leonardo da Vinci rule. For the same order fine root in different habitats, its length and specific surface area gradually increased from the edge of the forest to the canopy. The N content decreased first and then increased, while the C content and C/N increased first and then decreased. From the forest edge to the gap and to the under canopy, roots tended to move from the dichotomous branch to the herringbone-like branch by reducing the overlap between the secondary branches and roots, increasing specific root length, specific surface area and changing the contents of C and N to cope with environmental change and improve nutrient absorption efficiency.

Carnosic Acid Reverses the Inhibition of ApoE4 on Cell Surface Level of ApoER2 and Reelin Signaling Pathway.

The cell surface level of apolipoprotein E receptor 2 (ApoER2) increases by cyclic transport of ApoER2 and then activates Reelin signaling pathway to exert neuroprotective function in AD. ApoER2 ligand Apolipoprotein E4 (ApoE4) inhibits the recycling of ApoER2 to the cell surface rendering neurons unresponsive to Reelin. Carnosic acid (CA) is proven to possess neuroprotective and neurotrophic functions in Alzheimer's disease (AD) mouse model. However, there are few reports about how ApoE4 impairs the recycling of ApoER2 and if CA can affect the cyclic transport of ApoER2. In this study, we demonstrated that ApoE4 attenuates the binding of sorting nexin 17 (SNX17) to ApoER2 and inhibits the recycling of ApoER2, resulting in decreased cell surface level of ApoER2. Further, we found that CA enhances the binding of SNX17 to ApoER2, counteracts the negative effects of ApoE4 on the cell surface level of ApoER2 to reverse the ApoE4-induced reduction in Reelin signaling activation by increasing the phosphorylation of the N-methyl-D-aspartate receptor (NMDAR) and cAMP-response element-binding protein (CREB) and the expression of Gria2. Thus, CA promotes neurite growth inhibited by ApoE4. Our work suggests that CA may be a potential approach to attenuate the risk of ApoE4-associated AD.

SKP2 promotes breast cancer tumorigenesis and radiation tolerance through PDCD4 ubiquitination.

BACKGROUND: S-phase kinase-associated protein 2 (SKP2) is an oncogene and cell cycle regulator that specifically recognizes phosphorylated cell cycle regulator proteins and mediates their ubiquitination. Programmed cell death protein 4 (PDCD4) is a tumor suppressor gene that plays a role in cell apoptosis and DNA-damage response via interacting with eukaryotic initiation factor-4A (eIF4A) and P53. Previous research showed SKP2 may interact with PDCD4, however the relationship between SKP2 and PDCD4 is unclear. METHODS: To validate the interaction between SKP2 and PDCD4, mass spectrometric analysis and reciprocal co-immunoprecipitation (Co-IP) experiments were performed. SKP2 stably overexpressed or knockdown breast cancer cell lines were established and western blot was used to detect proteins changes before and after radiation. In vitro and in vivo experiments were performed to verify whether SKP2 inhibits cell apoptosis and promotes DNA-damage response via PDCD4 suppression. SMIP004 was used to test the effect of radiotherapy combined with SKP2 inhibitor. RESULTS: We found that SKP2 remarkably promoted PDCD4 phosphorylation, ubiquitination and degradation. SKP2 promoted cell proliferation, inhibited cell apoptosis and enhanced the response to DNA-damage via PDCD4 suppression in breast cancer. SKP2 and PDCD4 showed negative correlation in human breast cancer tissues. Radiotherapy combine with SKP2 inhibitor SMIP004 showed significant inhibitory effects on breast cancer cells in vitro and in vivo. CONCLUSIONS: We identify PDCD4 as an important ubiquitination substrate of SKP2. SKP2 promotes breast cancer tumorigenesis and radiation tolerance via PDCD4 degradation. Radiotherapy combine with SKP2-targeted adjuvant therapy may improve breast cancer patient survival in clinical medicine.

[Soil denitrification rates in different type riparian zones in maoershan mountainous region of China].

Soil samples were collected from forested, clear-cut, and grassy riparian zones under forest background and from forested and barren riparian zones under cropland background in the Maoershan mountainous region of China. The samples were incubated in laboratory, and their denitrification potentials were determined by nitrate-deduction method. The results showed that under crop-land background, soil denitrification rate was the highest in forested riparian zone and the lowest in barren riparian zone, with the deduction rate of nitrate varied from 46.79%-91.13% and 15.64% -81.84%, respectively. Under forest background, soil denitrification rate decreased in the order of clearcut > forested > grassy riparian zone, with the deduction rate of nitrate being 42.06%-90.39%, 28.24%-85.73% and 21.44%-83.11%, respectively. The denitrification rate was higher in subsurface layer than in deeper layer, and the denitrification potential was limited by the available carbon and nitrate, being the greatest in the forested riparian zone under cropland background.

[Impact of soil cadmium contamination on chlorophyll fluorescence characters and biomass accumulation of four broad-leaved tree species seedlings].

In a greenhouse pot experiment,a gradient of soil Cd concentration was established by adding 0,50,100 and 200 mg CdCl2 x kg(-1) air-dried soil, and the seedlings of one-year-old Rosa davurica, Philadelphus schrenkii, Acer mandshurica and Lonicera maackii were transplanted. The measurement of the seedlings' chlorophyll fluorescence parameters and biomass accumulation after 70 days of transplanting showed that with increasing soil Cd content, all the test tree species had a decrease of Fv/Fm, phi(PSII), qP and biomass, but an increase of qN. A. mandshurica had the greatest responses, followed by P. schrenkii and L. maackii, and R. davurica. The tolerance of these tree species to soil Cd contamination followed the order of R. davurica > P. schrenkii and L. maackii > A. mandshurica.

[Lymphatic metastasis intensity of and lymphadenectomy for thoracic esophageal squamous cell carcinoma].

BACKGROUND & OBJECTIVE: The intensity of lymphatic metastasis consists of lymph node metastasis number (LMN) and lymph node metastasis ratio (LMR). LMR is the ratio of positive nodes to dissected nodes. LMN and LMR are 2 important prognostic factors of esophageal cancer, and are adopted in tumor staging. This study was to assess the lymphatic metastasis intensity of thoracic esophageal squamous cell carcinoma (ESCC), and explore the influential factors and lymphadenectomy pattern. METHODS: A total of 120 patients with ESCC had been operated in the Second Affiliated Hospital of Sun Yat-sen University from 1998 to 2000. The lymph nodes were dissected according to the mapping scheme of the American Thoracic Society (ATS) modified by Casson et al. RESULTS: A total of 2 631 lymph nodes were dissected with an average of 22 lymph nodes in each case. The cervical LMR was significantly higher in the upper thoracic ESCC than in the middle and lower thoracic ESCC (20.9% vs. 12.9% and 6.8%, P<0.05). The left gastric LMR was significantly higher in the lower thoracic ESCC than in the middle and upper thoracic ESCC (37.5% vs. 17.5% and 7.1%, P<0.05). Subcarinal metastatic lymph nodes were often found in the middle thoracic ESCC. T stage, histological differentiation, and circum wall involvement degree were correlated to lymphatic metastasis intensity (P<0.05); the length of esophageal lesion had no correlation to lymphatic metastasis intensity (P>0.05). The survival time of the patients received the right thoracic esophagectomy with 3-field lymphadenectomy (3-FL) was significantly longer than that of the patients received the left thoracic esophagectomy with 2-FL (P<0.05). CONCLUSIONS: During the operation on ESCC, the regions with high lymphatic metastasis intensity should be dissected. T stage, histological differentiation, and circum wall involvement degree are important influential factors of lymphatic metastasis intensity. The right thoracic esophagectomy with 3-FL is superior to the left thoracic esophagectomy with 2-FL.