Dear Editor, N6-methyladenosine （m6A） has been demonstrated to be ubiquitous in several types of eukaryotic RNAs, including messenger RNA （mRNA）, transfer RNA （tRNA）, ribosomal RNA （rRNA）, long non-coding RNA （lncRNA）, and small nuclear RNA （snRNA） . The recent discoveries of RNA m6A methyltransferase complex METTL3/METTL14/WTAP and demethylases FTO and ALKBH5 prove the reversibility of m6A modification [2-6]. This modification plays important roles in various biological processes, including circadian rhythms ,
Aggressive NK-cell leukemia （ANKL） is a rare form of NK cell neoplasm that is more prevalent among people from Asia and Central and South America. Patients usually die within days to months, even after receiving prompt therapeutic management. Here we performed the first comprehensive study of ANKL by integrating whole genome, transcriptome and targeted sequencing, cytokine array as well as functional assays. Mutations in the JAK-STAT pathway were identified in 48% （14/29） of ANKL patients, while the extracelinlar STAT3 stimulator IL10 was elevated by an average of 56-fold （P 〈 0.0001） in the plasma of all patients examined. Additional frequently mutated genes included TP53 （34%）, TET2 （28%）, CREBBP （21%） and MLL2 （21%）. Patient NK leukemia cells showed prominent activation of STAT3 phosphorylation, MYC expression and transcriptional activities in multiple metabolic path- ways. Functionally, STAT3 activation and MYC expression were critical for the proliferation and survival of ANKL cells. STAT signaling regulated the MYC transcription program, and both STAT signaling and MYC transcription were required to maintain the activation of nucleotide synthesis and glycolysis, Collectively, the JAK-STAT path- way represents a major target for genomic alterations and IL10 stimulation in ANKL. This newly discovered JAK/ STAT-MYC-biosynthesis axis may provide opportunities for the development of novel therapeutic strategies in treating this subtype of leukemia.
Intestinal stem cells （ISCs） in the Drosophila adult midgut are essential for maintaining tissue homeostasis, and their proliferation and differentiation speed up in order to meet the demand for replenishing the lost cells in response to injury. Several signaling pathways including JAK-STAT, EGFR and Hippo （Hpo） pathways have been implicated in damage-induced ISC proliferation, but the mechanisms that integrate these pathways have remained elusive. Here, we demonstrate that the Drosophila homolog of the oncoprotein Myc （dMyc） functions downstream of these signaling pathways to mediate their effects on ISC proliferation, dMyc expression in precursor cells is stimulated in response to tissue damage, and dMyc is essential for accelerated ISC proliferation and midgut regeneration. We show that tissue damage caused by dextran sulfate sodium feeding stimulates dMyc expression via the Hpo pathway, whereas bleomycin feeding activates dMyc through the JAK-STAT and EGFR pathways. We provide evidence that dMyc expression is transcriptionally upregulated by multiple signaling pathways, which is required for optimal ISC proliferation in response to tissue damage. We have also obtained evidence that tissue damage can upregulate dMyc expression post-transcriptionally. Finally, we show that a basal level of dMyc expression is required for ISC mainte- nance, proliferation and lineage differentiation during normal tissue homeostasis.
Dear Editor, Cilia are remarkable microtubule （MT）-based organelles that are essential for cell motility, sensory perception and signal transduction . Ciliary defects have been implicated in various human diseases . Although the rod-shaped cilium morphology and the axonemal structure characterized by nine-doublet MTs were described decades ago [2, 3], the molecular regulations of cilium morphology and axonemal structure are still poorly defined.
The role of the hypoxia-inducible factor (HIF) subunits 1 alpha and 2 alpha in response to hypoxia. is well established in lung epithelial cells, whereas little is known about HIF-3 alpha with respect to transcriptional and translational regulation by hypoxia. HIF-3 alpha and HIF-1 alpha are two similar but distinct basic helix-loop-helix-PAS proteins, which have been postulated to activate hypoxia responsive genes in response to hypoxia. Here, we used quantitative real time RT-PCR and immunoblotting to determine the activation of HIF-3 alpha vs. HIF-1 alpha by hypoxia. HIF-3 alpha was strongly induced by hypoxia (1% O-2) both at the level of protein and mRNA due to an increase in protein stability and transcriptional activation, whereas HIF-1 alpha protein and mRNA levels enhanced transiently and then decreased because of a reduction in its mRNA stability in A549 cells, as measured on mRNA and protein levels. Interestingly, HIF-3 alpha and HIF-1 alpha exhibited strikingly similar responses to a variety of activating or inhibitory pharmacological agents. These results demonstrate that HIF-3 alpha is expressed abundantly in lung epithelial cells, and that the transcriptional induction of HIF-3 alpha plays an important role in the response to hypoxia in vitro. Our findings suggest that HIF-3 alpha, as a member of the HIF system, is complementary rather than redundant to HIF-1 alpha induction in protection against hypoxic damage, in alveolar epithelial cells.
Dear Editor, Recent discoveries suggest that N6-methyladenosine （m6A） modification, a prevalent internal modification in eukaryotic RNA, is an essential RNA regulatory mecha- nism. This modification is post-transcriptionally installed by m6A methyltransferases （METTL3-METTL14-WTAP complex） [1-4] and oxidatively removed by m6A demethylases （FTO and ALKBH5） [5, 6]. These ＇writer＇ and ＇eraser＇ enzymes are required for embryo development, energy homeostasis and fertility, suggesting fundamental regulatory roles of m6A [1, 2, 5].
AMP-activated protein kinase （AMPK） is a central cellular energy sensor and regulator of energy homeostasis, and a promising drug target for the treatment of diabetes, obesity, and cancer. Here we present low-resolution crystal structures of the human α1β2γ1 holo-AMPK complex bound to its allosteric modulators AMP and the glycogen-mimic cyclodextrin, both in the phosphorylated （4.05A^°） and non-phosphorylated （4.60A^°） state. In addition, we have solved a 2.95 structure of the human kinase domain （KD） bound to the adjacent autoinhibitory domain （AID） and have performed extensive biochemical and mutational studies. Together, these studies illustrate an underlying mechanism of allosteric AMPK modulation by AMP and glycogen, whose binding changes the equilibria between alternate AID （AMP） and carbohydrate-binding module （glycogen） interactions.
Protein ubiquitination by E3 ubiquitin ligases plays an important role in cancer development. In this study, we provide experimental evidence that a RING-finger-containing protein RNF13 is an ER/Golgi membrane-associated E3 ubiquitin ligase and its RING finger domain is required for the ubiquitin ligase activity. Immunohistochemical analysis of pancreatic ductal adenocarcinoma (PDAC) and paracancerous normal tissues from 72 patients documented RNF13 over-expression in 30 tumor samples (41.7%, 30/72), and its expression was significantly associated with histological grading (P = 0.024). In addition, RNF13 was detected in precancerous lesions: tubular complexes in chronic pancreatitis (CP) and pancreatic intraepithelial neoplasia (PanIN) (79.3%, 23/29 and 62.8%, 22/35, respectively). Moreover, RNF13 staining was significantly correlated with Tenascin-C expression (P = 0.004) in PDAC samples, further supporting the role of RNF13 in cancer progression. Over-expression of wild type but not RING domain-mutant RNF13 in pancreatic MiaPaca-2 cancer cells increased invasive potential and gelatinolytic activity by matrix metalloproteinase-9. Taken together, these findings reveal that RNF13 is a novel E3 ubiquitin ligase involved in pancreatic carcinogenesis; ubiquitin-mediated modification of proteins by RNF13 may participate in pancreatic cancer development.
Neurovascular interactions are crucial for the normal development of the central nervous system. To study such interactions in primary cultures, we developed a procedure to simultaneously isolate neural progenitor and endothelial cell fractions from embryonic mouse brains. Depending on the culture conditions endothelial cells were found to favor maintenance of the neuroprogenitor phenotype through the production of soluble factors, or to promote neuronal differentiation of neural progenitors through direct contact. These apparently opposing effects could reflect differential cellular interactions needed for the proper development of the brain.
Dear Editor, PIWI-interacting RNAs （piRNAs） are germ cell-specific small non-coding RNAs that are essential for silenc- ing transposable elements. Substantial efforts in the past decade have led to an understanding of how piRNAs are made. Primary piRNA biogenesis is initiated with transcription of piRNA precursors, followed by cleavage into piRNA intermediates, and finally, maturation by 3＇ end trimming and 2＇-O-methylation. Secondary piRNA biogenesis occurs through an amplification loop （ping pong pathway）; the piRNA pools generated through pri- mary processing guide MILI protein to cleave the target RNA for piRNA generation in a feed-forward loop that accelerates production of the piRNAs. Papi/Tdrkh has been implicated in processing the 3＇ ends of piRNAs [1, 2], however,