Related studies showed that the protein PSF represses protooncogene transcription, and VL30 -1 RNA, a mouse noncoding retroelement RNA, binds and releases PSF from a proto-oncogene, activating transcription. Here we show that this mechanism regulates tumorigenesis in human cells, with human RNAs replacing VL30 -1 RNA. A library of human RNA fragments was used to isolate, by affinity chromatography, 5 noncoding RNA fragments that bind to human PSF (hPSF), releasing hPSF from a protooncogene and activating transcription. T he protein PSF (1) contains a DNA-binding domain (DBD) that binds to the regulatory region of a proto-oncogene and represses transcription, and 2 RNA-binding domains (RBDs) that bind VL30-1 RNA, releasing PSF from a repressed protooncogene and activating transcription (2-5). Mouse and human genomes encode homologous PSF proteins with Ϸ95% sequence identity, whereas the VL30-1 gene belongs to a family of mouse noncoding retroelement genes (6) that is not present in the human genome (7). To determine whether the PSF/RNA regulatory mechanism functions in human cells, a library of RNA fragments was constructed from the nuclear RNA repertoire of a human tumor cell, and the library was screened by affinity chromatography for RNAs that bind to human PSF (hPSF). The screen identified 5 hPSF-binding noncoding RNA fragments that release hPSF from a repressed proto-oncogene and activate transcription, similar to VL30-1 RNA. Each human RNA fragment maps to a matching sequence in a different human gene. The following experiments show that human hPSF-binding RNAs are involved in the control of tumorigenesis. Results Cloning and Mapping Human RNA Fragments That Bind to hPSFProtein. The finding that VL30-1 RNA, a mouse retroelement RNA that is not encoded in the human genome, binds selectively to hPSF protein and reverses repression of proto-oncogene transcription (2-5), prompted a search for human RNAs that have a similar function as VL30-1 RNA. The procedure involved synthesizing a library of RNA fragments from the nuclear RNA repertoire of a human melanoma line and selecting by affinity chromatography RNA fragments that bind to hPSF. The procedure yielded 5 such RNA fragments, 4 of which were mapped, by sequence identity, within 1 of the following genes: L1PA16, a non-LTR retroelement gene (8); MER11C, a LTR retroelement gene (9); MALAT-1, a noncoding gene (10, 11); or HN, a mitochondrial gene coding for the peptide humanin (12); a fifth fragment, not shown in the figure, maps to a region that has not been characterized ( Fig. 1 and SI Text).The sequence of the HN RNA fragment is 100% identical to a sequence in the mitochondrial 16S ribosomal RNA gene and is 85% identical to positions 21947595-21947823 on nuclear chromosome 17. Further testing showed that the HN RNA fragment is derived from the mitochondrial HN RNA and not from the nuclear RNA (SI Text). The mitochondrial HN RNA might be translocated to the nucleus or derived from a mitochondrial contamination in the nuclear preparation.Release of hPSF from ...

SummaryMaize is a globally important food, feed crop and raw material for the food and energy industry. Plant architecture optimization plays important roles in maize yield improvement. PIN‐FORMED (PIN) proteins are important for regulating auxin spatiotemporal asymmetric distribution in multiple plant developmental processes. In this study, ZmPIN1a overexpression in maize increased the number of lateral roots and inhibited their elongation, forming a developed root system with longer seminal roots and denser lateral roots. ZmPIN1a overexpression reduced plant height, internode length and ear height. This modification of the maize phenotype increased the yield under high‐density cultivation conditions, and the developed root system improved plant resistance to drought, lodging and a low‐phosphate environment. IAA concentration, transport capacity determination and application of external IAA indicated that ZmPIN1a overexpression led to increased IAA transport from shoot to root. The increase in auxin in the root enabled the plant to allocate more carbohydrates to the roots, enhanced the growth of the root and improved plant resistance to environmental stress. These findings demonstrate that maize plant architecture can be improved by root breeding to create an ideal phenotype for further yield increases.

cancer therapy ͉ Rab23 ͉ regulatory RNA ͉ tumor suppression P receding studies have described a reversible mechanism controlling gene transcription that involves PSF protein (1) and VL30-1 RNA, a member of the VL30 family of mouse retroelement noncoding RNAs (2, 3). PSF contains a DNAbinding domain (DBD) that binds and represses transcription of genes that have a PSF-binding site (4-6) and 2 RNA-binding domains (RBDs) that bind VL30-1 RNA, forming a PSF-RNA complex that dissociates from a gene and activates transcription (5-7). Increasing expression of PSF in a human tumor cell suppressed tumorigenesis (6), and ectopic expression of VL30-1 RNA in a human tumor cell promoted metastasis (3). VL30 RNAs are expressed in virtually all tissues of adult mice (8) and are associated with Ras-mediated transformation of mouse fibroblast cells (9). Here, we extend our studies of the function of PSF and VL30-1 RNA to the regulation of proto-oncogene transcription, cell proliferation, and tumorigenesis in mice. The results indicate that PSF is a major tumor-suppressor protein and VL30-1 RNA is a major tumor-promoter RNA in mice. ResultsExpression of PSF and VL30 -1 RNA in NIH/3T3 and B16F10 Cell Lines.NIH/3T3 and B16F10 cells were transfected with a transgene encoding PSF (NIH/3T3-PSF1 and B16F10-PSF1 lines) or VL30-1 RNA (NIH/3T3-VL301 line) or with a plasmid encoding a shRNA that causes degradation of PSF mRNA (NIH/ 3T3-PSF2 line) or VL30 -1 RNA (NIH/3T3-VL302 and B16F10-VL302 lines). The control lines were transfected with an empty plasmid pcDNA3.1(ϩ) (NIH/3T3-pcDNA3.1 and B16F10-pcDNA3.1 cell lines) or a plasmid encoding shLuciferase (NIH/3T3-shLuc and B16F10-shLuc lines). The NIH/ 3T3 and B16F10 cell lines were assayed for expression of PSF mRNA and VL30-1 RNA ( Fig. 1 and Table 1) and PSF protein (Fig. 1).Binding of PSF to the Regulatory DNAs of Mouse Genes. Chromatin fragments from NIH/3T3 cells were coimmunoprecipitated with anti-PSF antibody, and the DNAs in the fragments were tested for hybridization to a mouse gene-promoter chip (NimbleGen; Roche) that contains 385,000 regulatory DNAs from the mouse genome. A total of 57 DNA fragments were identified by chip assay, of which 12 mapped to characterized mouse coding genes [supporting information (SI) Tables S1 and S2]. The 12 DNAs bound to PSF in NIH/3T3 and B16F10 cells, as shown by a ChIP assay (Fig. 2); the P450scc gene had been identified as a PSF-binding gene in earlier studies (4, 6). We chose the Rab23 gene, a member the RAS gene family (10,11), to analyze the role of PSF and VL30-1 RNA in the regulation of proto-oncogene transcription in mice. (Table 1). Binding of Rab23 DNA to PSF is higher in NIH/3T3-PSF1 and B16F10-PSF1 cells and lower in NIH/3T3-PSF2 cells than in NIH/3T3 or B16F10 control cells (Fig. 3A). (ii) Transcription of Rab23 was analyzed in the same cell lines by RT-PCR. Transcription is lower in NIH/3T3-PSF1 and B16F10-PSF1 cells and higher in NIH/3T3-PSF2 cells than in NIH/3T3 or B16F10 control cells (Fig. 3B). The results indicate that PSF bin...

Recombinant human growth differentiation factor 15 (rhGDF15) affects dendritic cell (DC) maturation. However, whether GDF15 is expressed in DCs and its roles and signaling in DCs remain largely unknown. It is unclear whether GDF15-DCs can induce immune tolerance in heart transplantation (HT). This study aims to understand the impact of endogenous GDF15 on DC's development, function, underlying molecular mechanism including circular RNA (circRNA). This study will also explore GDF15-DC-mediated immune modulation in HT. Bone marrow (BM) derived DCs were cultured and treated to up- or down regulate GDF15 expression. Phenotype and function of DCs were detected. Expression of genes and circRNAs was determined by qRT-PCR. The signaling pathways activated by GDF15 were examined. The impact of GDF15 treated DCs on preventing allograft immune rejection was assessed in a MHC full mismatch mouse HT model. Our results showed that GDF15 was expressed in DCs. Knockout of GDF15 promoted DC maturation, enhanced immune responsive functions, up-regulated malat-1 circular RNA (circ_Malat 1), and activated the nuclear factor kappa B (NFκB) pathway. Overexpression of GDF15 in DCs increased immunosuppressive/inhibitory molecules, enhanced DCs to induce T cell exhaustion, and promoted Treg generation through IDO signaling. GDF15 utilized transforming growth factor (TGF) β receptors I and II, not GFAL. Administration of GDF15 treated DCs prevented allograft rejection and induced immune tolerance in transplantation. In conclusion, GDF15 induces tolerogenic DCs (Tol-DCs) through inhibition of circ_Malat-1 and the NFκB signaling pathway and up-regulation of IDO. GDF15-DCs can prevent alloimmune rejection in HT.