Somatic mutations in the spliceosome gene ZRSR2—located on the X chromosome—are associated with myelodysplastic syndrome (MDS). ZRSR2 is involved in the recognition of 3′-splice site during the early stages of spliceosome assembly; however, its precise role in RNA splicing has remained unclear. Here we characterize ZRSR2 as an essential component of the minor spliceosome (U12 dependent) assembly. shRNA-mediated knockdown of ZRSR2 leads to impaired splicing of the U12-type introns and RNA-sequencing of MDS bone marrow reveals that loss of ZRSR2 activity causes increased mis-splicing. These splicing defects involve retention of the U12-type introns, while splicing of the U2-type introns remain mostly unaffected. ZRSR2-deficient cells also exhibit reduced proliferation potential and distinct alterations in myeloid and erythroid differentiation in vitro. These data identify a specific role for ZRSR2 in RNA splicing and highlight dysregulated splicing of U12-type introns as a characteristic feature of ZRSR2 mutations in MDS.

The Wnt signaling pathway is necessary both for maintaining undifferentiated stem cells and for directing their differentiation. In mouse embryonic stem cells (ESCs), Wnt signaling preferentially maintains “stemness” under certain permissive conditions. T-cell factor 3 (Tcf3) is a component of the Wnt signaling and a dominant downstream effector in ESCs. Despite the wealth of knowledge regarding the importance of Wnt signaling underlying stem cells functions, the precise mechanistic explanation by which the effects are mediated is unknown. In this study, we identified new regulatory targets of Tcf3 using a whole-genome approach and found that Tcf3 transcriptionally represses many genes important for maintaining pluripotency and self-renewal, as well as those involved in lineage commitment and stem cell differentiation. This effect is in part mediated by the corepressors transducin-like enhancer of split 2 and C-terminal Binding Protein (CtBP). Notably, Tcf3 binds to and represses the Oct4 promoter, and this repressive effect requires both the Groucho and CtBP interacting domains of Tcf3. Interestingly, we find that in mouse preimplantation development embryos, Tcf3 expression is coregulated with Oct4 and Nanog and becomes localized to the inner cell mass of the blastocyst. These data demonstrate an important role for Tcf3 in modulating the appropriate level of gene transcription in ESCs and during embryonic development. Disclosure of potential conflicts of interest is found at the end of this article.

Circular RNAs (circRNAs) are produced by head-to-tail back-splicing which is mainly facilitated by base-pairing of reverse complementary matches (RCMs) in circRNA flanking introns. Adenosine deaminases acting on RNA (ADARs) are known to bind double-stranded RNAs for adenosine to inosine (A-to-I) RNA editing. Here we characterize ADARs as potent regulators of circular transcriptome by identifying over a thousand of circRNAs regulated by ADARs in a bidirectional manner through and beyond their editing function. We find that editing can stabilize or destabilize secondary structures formed between RCMs via correcting A:C mismatches to I(G)-C pairs or creating I(G).U wobble pairs, respectively. We provide experimental evidence that editing also favors the binding of RNA-binding proteins such as PTBP1 to regulate back-splicing. These ADARs-regulated circRNAs which are ubiquitously expressed in multiple types of cancers, demonstrate high functional relevance to cancer. Our findings support a hitherto unappreciated bidirectional regulation of circular transcriptome by ADARs and highlight the complexity of cross-talk in RNA processing and its contributions to tumorigenesis.

Current standard of care for patients with pediatric acute lymphoblastic leukemia (ALL) is mainly effective, with high remission rates after treatment. However, the genetic perturbations that give rise to this disease remain largely undefined, limiting the ability to address resistant tumors or develop less toxic targeted therapies. Here, we report the use of next-generation sequencing to interrogate the genetic and pathogenic mechanisms of 240 pediatric ALL cases with their matched remission samples. Commonly mutated genes fell into several categories, including RAS/receptor tyrosine kinases, epigenetic regulators, transcription factors involved in lineage commitment, and the p53/cell-cycle pathway. Unique recurrent mutational hotspots were observed in epigenetic regulators CREBBP (R1446C/H), WHSC1 (E1099K), and the tyrosine kinase FLT3 (K663R, N676K). The mutant WHSC1 was established as a gain-of-function oncogene, while the epigenetic regulator ARID1A and transcription factor CTCF were functionally identified as potential tumor suppressors. Analysis of 28 diagnosis/relapse trio patients plus 10 relapse cases revealed four evolutionary paths and uncovered the ordering of acquisition of mutations in these patients. This study provides a detailed mutational portrait of pediatric ALL and gives insights into the molecular pathogenesis of this disease. Cancer Res; 77(2); 390–400. ©2016 AACR.

The nonimmune roles of Tregs have been described in various tissues, including the BM. In this study, we comprehensively phenotyped marrow Tregs, elucidating their key features and tissue-specific functions. We show that marrow Tregs are migratory and home back to the marrow. For trafficking, marrow Tregs use S1P gradients, and disruption of this axis allows for specific targeting of the marrow Treg pool. Following Treg depletion, the function and phenotype of both mesenchymal stromal cells (MSCs) and hematopoietic stem cells (HSCs) was impaired. Transplantation also revealed that a Treg-depleted niche has a reduced capacity to support hematopoiesis. Finally, we found that marrow Tregs are high producers of IL-10 and that Treg-secreted IL-10 has direct effects on MSC function. This is the first report to our knowledge revealing that Treg-secreted IL-10 is necessary for stromal cell maintenance, and our work outlines an alternative mechanism by which this cytokine regulates hematopoiesis.

RNA-binding proteins (RBPs) are key regulators of posttranscriptional processes such as RNA maturation, localization, turnover and translation. Despite their dysregulation in various diseases including cancer, the landscape of RBP expression in human cancer has not been well elucidated. Here, we built a comprehensive expression landscape of 1504 RBPs across 16 human cancer types, which revealed that RBPs are predominantly upregulated in tumours and this phenomenon is affected by the tumour immune subtypes and microenvironment. Across different cancer types, 109 RBPs are consistently upregulated while 41 RBPs are consistently downregulated. These up-regulated and down-regulated RBPs show distinct molecular characteristics and prognostic effects, whereas their dysregulation is mediated by distinct cis/trans-regulatory mechanisms. Finally, we validated one candidate PABPC1L that might promote colon tumorigenesis by regulating mRNA splicing. In summary, we built a comprehensive expression landscape of RBPs across different cancer types and identified consistently dysregulated RBPs which could be novel targets for developing broad-spectrum anticancer agents.