In contrast to cytoplasmic localization of spliced mRNAs, many spliced lncRNAs are localized in the nucleus. To investigate the mechanism, we used lncRNA MEG3 as a reporter and mapped a potent nuclear retention element (NRE), deletion of this element led to striking export of MEG3 from the nucleus to the cytoplasm. Insertion of the NRE resulted in nuclear retention of spliced lncRNA as well as spliced mRNA. We further purified RNP assembled on the NRE in vitro and identified the proteins by mass spectrometry. Screen using siRNA revealed depletion of U1 snRNP components SNRPA, SNRNP70 or SNRPD2 caused significant cytoplasmic localization of MEG3 reporter transcripts. Co-knockdown these factors in HFF1 cells resulted in an increased cytoplasmic distribution of endogenous lncRNAs. Together, these data support a model that U1 snRNP components restrain spliced lncRNAs in the nucleus via the interaction with nuclear retention element.

Xeroderma pigmentosum group A (XPA)-binding protein 2 (XAB2) is a multi-functional protein that plays critical role in processes including transcription, transcription-coupled DNA repair, pre-mRNA splicing, homologous recombination and mRNA export. Microarray analysis on gene expression in XAB2 knockdown cells reveals that many genes with significant change in expression function in mitotic cell cycle regulation. Fluorescence-activated cell scanner analysis confirmed XAB2 depletion led to cell arrest in G2/M phase, mostly at prophase or prometaphase. Live cell imaging further disclosed that XAB2 knockdown induced severe mitotic defects including chromosome misalignment and defects in segregation, leading to mitotic arrest, mitotic catastrophe and subsequent cell death. Among top genes down-regulated by XAB2 depletion is mitotic motor protein centrosome-associated protein E (CENPE). Knockdown CENPE showed similar phenotypes to loss of XAB2, but CENPE knockdown followed by XAB2 depletion did not further enhance cell cycle arrest. Luciferase assay on CENPE promoter showed that overexpression of XAB2 increased luciferase activity, whereas XAB2 depletion resulted in striking reduction of luciferase activity. Further mapping revealed a region in CENPE promoter that is required for the transcriptional regulation by XAB2. Moreover, ChIP assay showed that XAB2 interacted with CENPE promoter. Together, these results support a novel function of XAB2 in mitotic cell cycle regulation, which is partially mediated by transcription regulation on CENPE.

Photoperiod plays a key role in controlling the phase transition from vegetative to reproductive growth in flowering plants. Leaves are the major organs perceiving day-length signals, but how specific leaf cell-types respond to photoperiod remains unknown. We integrated photoperiod-responsive chromatin accessibility and transcriptome data in leaf epidermis and vascular companion cells of Arabidopsis thaliana by combining INTACT (Isolation of Nuclei Tagged in specific Cell/Tissue types) with ATAC-seq (Assay for Transposase-Accessible Chromatin using sequencing) and RNA-sequencing. Despite a large overlap, vasculature and epidermis cells responded differently. Long-day predominantly induced accessible chromatin regions (ACRs); in the vasculature, more ACRs were induced and these were located at more distal gene regions, compared with the epidermis. Vascular ACRs induced by long day were highly enriched in binding sites for flowering-related transcription factors. Among the highly ranked genes (based on chromatin and expression signatures in the vasculature), we identified TREHALOSE-6-PHOSPHATASE SYNTHASE 9 (TPS9) as a flowering activator, as shown by the late flowering phenotypes of T-DNA insertion mutants and transgenic lines with phloem-specific knockdown of TPS9. Our cell-type-specific analysis sheds light on how the long-day photoperiod stimulus impacts chromatin accessibility in a tissue-specific manner to regulate plant development.

Objective We retrospectively studied the efficacy of personalized therapy with surgical resection plus prophylactic management of postsurgical auricular keloids by intralesional injection of betamethasone and local pressure therapy using magnets in patients with auricular keloids. Methods Surgical excision was performed in all patients, and surgical techniques including fusiform excision of the keloid scar, core excision of the keloid scar followed by flap repair, and scar graft were chosen. Results A total of 85 patients with 98 auricular keloids were eligible. Seventy‐two (74%) patients had primary auricular keloids, and 13 patients had recurrent keloids after surgical excision. Keloids, were located in the helix in 28 (32.9%) cases, in the earlobe in 45 (52.9%) cases, and in the entire auricle in 12 (14.1%) cases. The size of auricular keloids ranged from 10 to 35 mm. Surgical resection was uneventful in all cases. Twenty‐one (21.4%) patients received fusiform excision, 47 (47.9%) patients underwent core excision and flap repair, and 30 (30.6%) patients received skin grafts. The patients were followed up for median duration of 1 year (range: 12–24 months). The cure rate was 87.2%, and the recurrence rate was 12.8%. Conclusion A personalized surgical approach based on the characteristics of auricular keloids in each patient and a multimodal therapeutic regimen including surgical excision, glucocorticoid blockade, and intralesional injection of glucocorticoids and pressure therapy improve the cure rate and reduce the recurrence rate of auricular keloids.

Melanoma is a malignant tumor. The acquisition of stemness by melanoma cells aggravates the malignant transformation, which can be regulated by microRNAs (miRNAs, miR). MiR-363-3p is a key tumor-related miRNA, but its role in stemness and melanoma cells is still unknown. Presently, miR-363-3p induced by hypoxia inducible factor (HIF)-2α has a positive role in melanoma cell stemness. The levels of miR-363-3p and HIF-2α are upregulated in melanoma cell lines. Overexpression of HIF-2α significantly increased levels of miR-363-3p. However, both HIF-2α knockdown and miR-363-3p inhibition decreased the levels of stemness markers (CD133, CD271, Jarid1B and Nanog). Furthermore, the levels of miR-363-3p and HIF-2α were upregulated in fluorescence activated cell sorting (FACS)-sorted CD271 high/+ cells. Whereas, miR-363-3p depletion reduced the proportion and the ability of the CD271 high/+ cells to form spheroids, decreased the levels of CD133, CD271, Jarid1B and Nanog with restrained proliferative activity of CD271 high/+ cells. Additionally, miR-363-3p was confirmed a key downstream effector of HIF-2α. Intriguingly, cyclin-dependent kinase inhibitor 1A [CDKN1A, p21(Cip1/Waf1)], a key inhibitor of S-phase DNA synthesis and cell cycle progression, was confirmed a target gene of miR-363-3p by luciferase reporter gene assay. The protein levels of CD133, CD271, Jarid1B and Nanog were upregulated with enhanced proliferative activity of CD271 high/+ cells by inhibition of p21 in melanoma cells. In conclusion, miR-363-3p is induced by HIF-2α to promote stemness of melanoma cells via inhibiting p21. The present study provides novel insights and indicates that HIF-2α/miR-363-3p/p21 signaling may be a potential target in melanoma research and therapy.