DNMT1 is recruited by PCNA and UHRF1 to maintain DNA methylation after replication. UHRF1 recognizes hemimethylated DNA substrates via the SRA domain, but also repressive H3K9me3 histone marks with its TTD. With systematic mutagenesis and functional assays, we could show that chromatin binding further involved UHRF1 PHD binding to unmodified H3R2. These complementation assays clearly demonstrated that the ubiquitin ligase activity of the UHRF1 RING domain is required for maintenance DNA methylation. Mass spectrometry of UHRF1-deficient cells revealed H3K18 as a novel ubiquitination target of UHRF1 in mammalian cells. With bioinformatics and mutational analyses, we identified a ubiquitin interacting motif (UIM) in the N-terminal regulatory domain of DNMT1 that binds to ubiquitinated H3 tails and is essential for DNA methylation in vivo. H3 ubiquitination and subsequent DNA methylation required UHRF1 PHD binding to H3R2. These results show the manifold regulatory mechanisms controlling DNMT1 activity that require the reading and writing of epigenetic marks by UHRF1 and illustrate the multifaceted interplay between DNA and histone modifications. The identification and functional characterization of the DNMT1 UIM suggests a novel regulatory principle and we speculate that histone H2AK119 ubiquitination might also lead to UIM-dependent recruitment of DNMT1 and DNA methylation beyond classic maintenance.
DNA methylation is an important epigenetic mark involved in diverse biological processes. In plants, DNA methylation can be established through the RNA-directed DNA methylation pathway, an RNA interference pathway for transcriptional gene silencing （TGS）, which requires 24-nt small interfering RNAs. In mammals, de novo DNA methylation occurs primarily at two developmental stages： during early embryogenesis and during gametogenesis. While it is not clear whether establishment of DNA methylation patterns in mammals involves RNA interference in general, de novo DNA methylation and suppression of transposons in germ cells require 24-32-nt piwi-interacting small RNAs. DNA methylation status is dynamically regulated by DNA methylation and demethylation reactions. In plants, active DNA demethylation relies on the repressor of silencing 1 family of bifunctional DNA glycosylases, which remove the 5-methylcytosine base and then cleave the DNA backbone at the abasic site, initiating a base excision repair （BER） pathway. In animals, multiple mechanisms of active DNA demethylation have been proposed, including a deaminase- and DNA glycosylase-initiated BER pathway. New information concerning the effects of various histone modifications on the establishment and maintenance of DNA methylation has broadened our understanding of the regulation of DNA methylation. The function of DNA methylation in plants and animals is also discussed in this review.
Two distinct nuclear factor kappa B (NF kappa B) signaling pathways have been described; the canonical pathway that mediates inflammatory responses, and the non-canonical pathway that is involved in immune cell differentiation and maturation and secondary lymphoid organogenesis. The former is dependent on the I kappa B kinase adaptor molecule NEMO, the latter is independent of it. Here, we review the molecular mechanisms of regulation in each signaling axis and attempt to relate the apparent regulatory logic to the physiological function. Further, we review the recent evidence for extensive cross-regulation between these two signaling axes and summarize them in a wiring diagram. These observations suggest that NEMO-dependent and -independent signaling should be viewed within the context of a single NF kappa B signaling system, which mediates signaling from both inflammatory and organogenic stimuli in an integrated manner. As in other regulatory biological systems, a systems approach including mathematical models that include quantitative and kinetic information will be necessary to characterize the network properties that mediate physiological function, and that may break down to cause or contribute to pathology.
Despite advances in DNA methylome analyses of cells and tissues, current techniques for genome-scale profiling of DNA methylation in circulating cell-free DNA （ccfDNA） remain limited. Here we describe a methylated CpG tan- dems amplification and sequencing （MCTA-Seq） method that can detect thousands of hypermethylated CpG islands simultaneously in ccfDNA. This highly sensitive technique can work with genomic DNA as little as 7.5 pg, which is equivalent to 2.5 copies of the haploid genome. We have analyzed a cohort of tissue and plasma samples （n = 151） of hepatocellular carcinoma （HCC） patients and control subjects, identifying dozens of high-performance markers in blood for detecting smaU HCC （≤ 3 cm）. Among these markers, 4 （RGS10, ST8SIA6, RUNX2 and VIM） are mostly specific for cancer detection, while the other 15, classified as a novel set, are already hypermethylated in the normal liver tissues. Two corresponding classifiers have been established, combination of which achieves a sensitivity of 94% with a specificity of 89% for the plasma samples from HCC patients （n = 36） and control subjects including cirrho- sis patients （n = 17） and normal individuals （n = 38）. Notably, all 15 alpha-fetoprotein-negative HCC patients were successfully identified. Comparison between matched plasma and tissue samples indicates that both the cancer and noncancerous tissues contribute to elevation of the methylation markers in plasma. MCTA-Seq will facilitate the development of ccfDNA methylation biomarkers and contribute to the improvement of cancer detection in a clinical setting.
MicroRNA （miRNA） biogenesis is finely controlled by complex layers of post-transcriptional regulators, including RNA-binding proteins （RBPs）. Here, we show that an RBP, QKI5, activates the processing of primary miR-124- 1 （pri-124-1） during erythropoiesis. QKI5 recognizes a distal QKI response element and recruits Microprocessor through interaction with DGCRS. Furthermore, the recruited Microprocessor is brought to pri-124-1 stem loops by a spatial RNA-RNA interaction between two complementary sequences. Thus, mutations disrupting their base-pairing affect the strength of QKI5 activation. When erythropoiesis proceeds, the concomitant decrease of QKI5 releases Mi- croprocessor from pri-124-1 and reduces mature miR-124 levels to facilitate erythrocyte maturation. Mechanistically, miR-124 targets TALl and c-MYB, two transcription factors involved in normal erythropoiesis. Importantly, this QKI5-mediated regulation also gives rise to a unique miRNA signature, which is required for erythroid differentiation. Taken together, these results demonstrate the pivotal role of QKI5 in primary miRNA processing during erythropoiesis and provide new insights into how a distal element on primary transcripts affects miRNA biogenesis.
New gene origination is a major source of genomic innovations that confer phenotyp ic changes and biological di- versity. Generation of new mitochondrial genes in plants may cause cytoplasmic male sterility （CMS）, which can pro- mote outcrossing and increase fitness. However, how mitochondrial genes originate and evolve in structure and func- tion remains unclear. The rice Wild Abortive type of CMS is conferred by the mitochondrial gene WA352c （previously named WA352） and has been widely exploited in hybrid rice breeding. Here, we reconstruct the evolutionary tra- jectory of WA352c by the identification and analyses of 11 mitochondrial genomic recombinant structures related to WA352c in wild and cultivated rice. We deduce that these structures arose through multiple rearrangements among conserved mitochondrial sequences in the mitochondrial genome of the wild rice Oryza rufipogon, coupled with sub- stoichiometric shifting and sequence variation. We identify two expressed but nonfunctional protogenes among these structures, and show that they could evolve into functional CMS genes via sequence variations that could relieve the self-inhibitory potential of the proteins. These sequence changes would endow the proteins the ability to interact with the nucleus-encoded mitochondrial protein COXI1, resulting in premature programmed cell death in the anther ta- petum and male sterility. Furthermore, we show that the sequences that encode the COXll-interaction domains in these WA352c-related genes have experienced purifying selection during evolution. We propose a model for the for- mation and evolution of new CMS genes via a ＂multi-recombination/protogene formation/functionalization＂ mecha- nism involving gradual variations in the structure, sequence, copy number, and function.
Maintaining protein homeostasis is essential for the survival of animals. A recent study in Cell Research identifies a group of DH44(+) neuroendocrine cells in the Drosophila brain as a novel fast-acting amino acid sensor, which detects three specific dietary amino acids and promotes food intake.
An imbalance of mucosal proand anti-inflammatory cytokincs is crucial in the pathogenesis of inflammatory bowel disease （IBD）. GM-CSF influences the development of hemopoietic cells. The precise role of GM-CSF in IBD remains to be elucidated. GM-CSF gene knockout （GM-CSF^-/-） and wild-type （Wt） mice were challenged with 2.5% dextran sulfate sodium （DSS） for 7 days. The ensued clinical and pathological changes, macrophage infiltration, colonic cytokine production, and bacterial counts were examined. DSS-treated GM-CSF^-/- mice developed more severe acute colitis than DSS-treated Wt mice, reflected by a greater body weight loss, more rectal bleeding, and aggravated histopathological changes. More infiltrating macrophages were observed in GM-CSF^-/-, compared with Wt mice following DSS challenge, correlating with monocyte chemoattractant protein-1 （MCP-1） production. The levels of colonic IL-17 and TNF-α were increased significantly in GM-CSF^-/- mice, but not in Wt mice, following DSS administration. The level of IL-6 was increased by 1.5- and 2-fold in the colon of GM-CSF^-/- and Wt mice, respectively, following DSS challenge. No significant changes in IL-4 and IFN-γ were detected in Wt and GM-CSF^-/- mice following DSS treatment. The bacteria recovery from colon was increased about 15- and 5-fold, respectively, in Wt mice and GM-CSF^-/- mice following DSS challenge. These results suggest that GM-CSF^-/- mice are more susceptible to acute DSS-induced colitis, possibly because of an impaired gut innate immune response as a result of diminished GM-CSF.
Dear Editor, Chronic virus infection, such as infection by Ep- stein-Barr virus （EBV）, hepatitis B/C virus （HBV/HCV） and human immunodeficiency virus （HIV）, constitutes major public health concerns. Although efforts in de- ciphering the mechanisms underlying their virological consequences have greatly improved clinical prevention and therapy, various challenges, such as virus tropism drifting, remain to be addressed to develop effective clin- ical interventions. While cell-free viruses usually infect target cells via binding to specific receptors, the presence of virions in non-susceptible cells has been reported with the mechanisms poorly understood .
Dear Editor, The midbody is a structure formed within the inter- cellular bridge towards the end of cytokinesis [ 1 ]. Microtubules within this bridge are then severed on one side of the midbody during abscission, thus generating a midbody remnant in one of the resulting daughter cells. Midbody remnants persist long after cell division and ac- cumulate preferentially in stem cells, induced pluripotent stem （iPS） cells, and cancer stem cells [2-4]. Upon in- duction of differentiation, midbody remnants are degrad- ed by autophagy or released into the extracellular milieu in some tissue culture cells, and it has been proposed that such removal is critical for enabling a differentiation pro- gram [2-4]. However, the fate of midbody remnants in a developing organism remains elusive, and whether their presence plays a role in cell fate determination in vivo is not known.
A classical voltage-gated ion channel consists of four voltage-sensing domains （VSDs）. However, the roles of each VSD in the channels remain elusive. We developed a GVTDT （Graft VSD To Dimeric TASK3 channels that lack endogenous VSDs） strategy to produce voltage-gated channels with a reduced number of VSDs. TASK3 channels exhibit a high host tolerance to VSDs of various voltage-gated ion channels without interfering with the intrinsic properties of the TASK3 selectivity filter. The constructed channels, exemplified by the channels grafted with one or two VSDs from Kv7.1 channels, exhibit classical voltage sensitivity, including voltage-dependent opening and closing. Furthermore, the grafted Kv7.1 VSD transfers the potentiation activity of benzbromarone, an activator that acts on the VSDs of the donor channels, to the constructed channels. Our study indicates that one VSD is sufficient to voltage-dependently gate the pore and provides new insight into the roles of VSDs.
The incidence of genetic material or epigenetic information transferred from one organism to another is an important biological question. A recent study demonstrated that plant small RNAs acquired orally through food intake directly influence gene expression in animals after migration through the plasma and delivery to specific organs.
Global shortening of 3＇ untranslated regions （3＇ UTRs） through alternative polyadenylation is an emerging hallmark of cancer. A recent study identifies the cleavage factor Im 25 （CFIm25） as an important mediator of 3＇ UTR shortening in glioblastomas and demonstrates a causal relationship between alternative polyadenylation and cancer cell proliferation.
Cells and organisms adapt to mitochondrial dysfunction by activating the mitochondrial unfolded protein response （UPRmt）, which is regulated by mitochondrial-to-nuclear communication; and UPRmt activation can also be transmitted between different cell types suggesting a role in tissue coordination. Shao and colleagues now identify a neuronal circuit and a secreted neuropeptide required for cell non-autonomous UPRmt regulation.