Initiation of zygotic transcription in mammals is poorly understood. In mice, zygotic transcription is first detected shortly after pronucleus formation in 1-cell embryos, but the identity of the transcribed loci and mechanisms regulating their expression are not known. Using total RNA-Seq, we have found that transcription in 1-cell embryos is highly promiscuous, such that intergenic regions are extensively expressed and thousands of genes are transcribed at comparably low levels. Striking is that transcription can occur in the absence of defined core-promoter elements. Furthermore, accumulation of translatable zygotic mRNAs is minimal in 1-cell embryos because of inefficient splicing and 3 0 processing of nascent transcripts. These findings provide novel insights into regulation of gene expression in 1-cell mouse embryos that may confer a protective mechanism against precocious gene expression that is the product of a relaxed chromatin structure present in 1-cell embryos. The results also suggest that the first zygotic transcription itself is an active component of chromatin remodeling in 1-cell embryos.

Lysine-specific demethylase 1 (LSD1) regulates gene expression by affecting histone modifications and is a promising target for acute myeloid leukemia (AML) with specific genetic abnormalities. Novel LSD1 inhibitors, NCD25 and NCD38, inhibited growth of MLL-AF9 leukemia as well as erythroleukemia, megakaryoblastic leukemia and myelodysplastic syndromes (MDSs) overt leukemia cells in the concentration range that normal hematopoiesis was spared. NCD25 and NCD38 invoked the myeloid development programs, hindered the MDS and AML oncogenic programs, and commonly upregulated 62 genes in several leukemia cells. NCD38 elevated H3K27ac level on enhancers of these LSD1 signature genes and newly activated ~500 super-enhancers. Upregulated genes with super-enhancer activation in erythroleukemia cells were enriched in leukocyte differentiation. Eleven genes including GFI1 and ERG, but not CEBPA, were identified as the LSD1 signature with super-enhancer activation. Super-enhancers of these genes were activated prior to induction of the transcripts and myeloid differentiation. Depletion of GFI1 attenuated myeloid differentiation by NCD38. Finally, a single administration of NCD38 causes the in vivo eradication of primary MDS-related leukemia cells with a complex karyotype. Together, NCD38 derepresses super-enhancers of hematopoietic regulators that are silenced abnormally by LSD1, attenuates leukemogenic programs and consequently exerts anti-leukemic effect against MDS-related leukemia with adverse outcome.

Human endogenous retroviruses (HERV) are associated with many diseases such as autoimmune diseases and cancer. Although the frequent expression of a variety of HERVs in tumor cells has been demonstrated, their functional contributions in cancer are as yet unclear. Intriguingly, HERVs and other retroviruses include an immunosuppressive domain in their transmembrane envelope proteins, but its mechanism of action and cancer relevance are obscure. In this study, we demonstrate that the human endogenous retrovirus HERV-H has a critical role in tumor metastasis and immune escape. We found that expression of herv-h mRNA was elevated in metastatic tumor cells undergoing epithelial-to-mesenchymal transition (EMT) and in primary tumor tissues from advanced colon cancer. The immunosuppressive peptide H17 derived from HERV-H was sufficient to induce EMT in tumor cells that expressed low levels of HERV-H, and it amplified this event within the tumor microenvironment. H17 also stimulated CCL19 expression in tumor cells, which in turn recruited and expanded a population of pluripotent immunoregulatory CD271 þ cells, which included mesenchymal stem cells and myeloid-derived suppressor cells. In tumor tissues from patients with advanced colon cancer, we confirmed that CD271 þ cells were increased in HERV-H þ CCL19 þ tumor tissues. Notably, RNAi-mediated change of HERV-H or CCL19, or depletion of CD271 þ cells, improved immune responses in vitro and in vivo accompanied by tumor regression. Together, our results argued that HERV-H is a critical determinant of immune escape in cancer, suggesting its candidacy as a promising therapeutic target to treat patients with advanced cancer.

BACKGROUND & AIMS: Proper resolution of inflammation is essential to maintaining homeostasis, which is important as a dysregulated inflammatory response has adverse consequences, even being regarded as a hallmark of cancer. However, our picture of dynamic changes during inflammation remains far from comprehensive. METHODS:Here we used single-cell transcriptomics to elucidate changes in distinct cell types and their interactions in a mouse model of chemically induced colitis.RESULTS: Our analysis highlights the stromal cell population of the colon functions as a hub with dynamically changing roles over time. Importantly, we found that Serpina3n, a serine protease inhibitor, is specifically expressed in stromal cell clusters as inflammation resolves, interacting with a potential target, elastase. Indeed, genetic ablation of the Serpina3n gene delays resolution of induced inflammation. Furthermore, systemic Serpina3n administration promoted the resolution of inflammation, ameliorating colitis symptoms. CONCLUSIONS:This study provides a comprehensive, single-cell understanding of cell-cell interactions during colorectal inflammation and reveals a potential therapeutic target that leverages inflammation resolution.

Mesenchymal stem cells (MSCs), which can differentiate into tri-lineage (osteoblast, adipocyte, and chondrocyte) and suppress inflammation, are promising tools for regenerative medicine. MSCs are phenotypically diverse based on their tissue origins. However, the mechanisms underlying cell-type-specific gene expression patterns are not fully understood due to the lack of suitable strategy to identify the diversity. In this study, we investigated gene expression programs and chromatin accessibilities of MSCs by whole-transcriptome RNA-seq analysis and an assay for transposase-accessible chromatin using sequencing (ATAC-seq). We isolated MSCs from four tissues (femoral and vertebral bone marrow, adipose tissue, and lung) and analysed their molecular signatures. RNA-seq identified the expression of MSC markers and both RNA-seq and ATAC-seq successfully clustered the MSCs based on their tissue origins. Interestingly, clustering based on tissue origin was more accurate with chromatin accessibility signatures than with transcriptome profiles. Furthermore, we identified transcription factors potentially involved in establishing cell-type specific chromatin structures. Thus, epigenome analysis is useful to analyse MSC identity and can be utilized to characterize these cells for clinical use.

Lysine-specific demethylase 1 (LSD1) is a histone modifier for transcriptional repression involved in the regulation of hematopoiesis. We previously reported that a LSD1 inhibitor NCD38 induces transdifferentiation from erythroid lineage to granulomonocytic lineage and exerts anti-leukemia effect through de-repression of the specific super-enhancers of hematopoietic regulators including ERG in a human erythroleukemia cell line, HEL. However, the mechanistic basis for this specificity of NCD38 has remained unclear. Herein, we report major partners associated with LSD1 and clarify the mechanism in HEL cells. Proteome analysis identified 54 candidate proteins associated with LSD1, including several transcription factors such as GFI1B and RUNX1 as well as BRAF-histone deacetylase complex (BHC) components such as CoREST, HDAC1, and HDAC2. NCD38 selectively disrupted the interaction of LSD1 with GFI1B but not with RUNX1, CoREST, HDAC1 and HDAC2. Erg was downregulated in murine erythroid progenitors with prominent upregulation of Gfi1b. NCD38 induced ERG and attenuated an erythroid marker CD235a in HEL while this attenuation was mimicked by the lentiviral overexpression of ERG. The ERG super-enhancer contained the conserved binding motif of GFI1B and was actually occupied by GFI1B. NCD38 dissociated LSD1 and CoREST but not GFI1B from the ERG super-enhancer. Collectively, the selective separation of LSD1 from GFI1B by NCD38 restores the ERG super-enhancer activation and consequently upregulates ERG expression, inducing the transdifferentiation linked to the anti-leukemia effect.

The objective of this study is to describe a technique for preserving the autonomic nerve systematically, including the hypogastric nerves, pelvic splanchnic nerves, and pelvic plexus and its vesical branches, based on anatomic considerations for the autonomic nerves innervating the urinary bladder, in radical hysterectomies and to assess postsurgical bladder function. A nerve-sparing radical hysterectomy was carried out on 27 consecutive patients with uterine cervical cancer treated between 2000 and 2002. The FIGO stages of the disease consisted of 10 stage Ib1, 6 stage Ib2, 3 stage IIa, and 8 stage IIb. The nerve-sparing procedure was successfully completed in 22 of the 27 patients (81.5%) in the study. At 1 year after the operation, bladder symptoms were significantly improved in the nerve-sparing group compared to the non–nerve-sparing group. Urinary incontinence and abnormal (diminished) bladder sensation were observed in three of the five patients (two patients had both symptoms), for whom the nerve-sparing procedure could not be performed, but none of the 22 patients for whom the nerve-sparing procedure was performed had incontinence, and only two patients had abnormal (increased) bladder sensation (P = 0.0034 for incontinence and P = 0.030 for abnormal bladder sensation). The patients' survival was not adversely affected by the nerve-sparing procedure. Although it is still preliminary, the surgical technique described in this report is thought to be effective for preserving bladder function, and thus, the quality of life could be improved for patients with cervical cancer who are treated with a radical hysterectomy. For further evaluation of the efficacy of nerve-sparing radical hysterectomy, a prospective randomized trial needs to be performed.

Bendamustine has rapidly become one of the key drugs for patients with indolent lymphoma because of its safety, tolerability and eff ectiveness. However, two of the 11 patients we have administered the drug developed severe immunosuppression against cytomegalovirus (CMV). Bendamustine potentially hampers immune function and should be used more cautiously.Case 1 A 51-year-old man was diagnosed with follicular lymphoma with massive bone marrow infi ltration in 2006. He received R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine and prednisolone) chemotherapy that resulted in complete remission, but underwent a few relapses, requiring a R-FND (rituximab, fl udarabine, mitoxantrone and dexamethasone) salvage regimen. At the fi fth relapse in 2011, the regimen was switched to cycles of 120 mg/m 2 /day bendamustine (days 1 -2) every 28 days. During the third cycle, the patient developed severe diarrhea, and lost more than 10 kg in weight. His white blood cell (WBC) count was 2.8 ϫ 10 9 /L with 93% neutrophils and 2.5% lymphocytes. Other relevant laboratory results were aspartate aminotransferase (AST) level, 111 U/L; alanine aminotransferase (ALT) level, 110 U/L; and immunoglobulin G (IgG) level, 339 mg/dL. Th e CD4 count was 21/ μ L with a CD4/8 ratio of 0.16. A positive result for CMV pp65 antigen C10/11 (603/689) and the presence of retinitis strongly suggested that the patient had an active CMV infection. Although the patient ' s serum briefl y became negative for the CMV pp65 antigen after intensive treatment with ganciclovir or foscarnet, a repeat of CMV-associated symptoms and a recurrence of CMV serum positivity were noted on discontinuation of anti-CMV therapy for more than 1 year. Furthermore, CMV-specifi c CD8 ϩ cytotoxic T lymphocytes (CTLs) were persistently undetectable in four samples taken every month using tetramer-based quantifi cation [1], indicating a profound and protracted immune defi ciency against CMV. Reduced intensity hematopoietic stem cell transplant from an unrelated donor was performed to reconstruct cellular immunity against CMV as well as to eradicate residual follicular lymphoma cells. On day 15, the patient suff ered acute renal failure, followed by sudden cardiac arrest. He was emergently resuscitated once, but fi nally died because of multi-organ failure presumably due to CMV-associated hemophagocytic syndrome on day 59 after transplant.Case 2 An 89-year-old woman was diagnosed with diff use large B-cell lymphoma. Despite the administration of two cycles of R-COP (rituximab, cyclophosphamide, vincristine and prednisolone) chemotherapy, only a small response was observed. She then received two cycles of bendamustine at the same dose as the patient in Case 1, because she appeared too frail to be treated by a doxorubicin-containing regimen such as CHOP chemotherapy. She developed nausea and appetite loss 10 days after the last administration of bendamustine. Her WBC count was 4.8 ϫ 10 9 /L with 89% neutrophils and 4.0% lymphocytes. Other relevant laboratory fi ndings were AST...