N 6-methyladenosine (m 6 A) is the most abundant mRNA modification and is catalyzed by the methyltransferase complex, in which methyltransferase-like 3 (METTL3) is the sole catalytic subunit. Accumulating evidence in recent years reveals that METTL3 plays key roles in a variety of cancer types, either dependent or independent on its m 6 A RNA methyltransferase activity. While the roles of m 6 A modifications in cancer have been extensively reviewed elsewhere, the critical functions of METTL3 in various types of cancer, as well as the potential targeting of METTL3 as cancer treatment, have not yet been highlighted. Here we summarize our current understanding both on the oncogenic and tumor-suppressive functions of METTL3, as well as the underlying molecular mechanisms. The welldocumented protein structure of the METTL3/METTL14 heterodimer provides the basis for potential therapeutic targeting, which is also discussed in this review.

N6-methyladenosine (m6A) is a reversible and dynamic RNA modification in eukaryotes. However, how cells establish cell-specific m6A methylomes is still poorly understood. Here, we developed a computational framework to systematically identify cell-specific trans regulators of m6A through integrating gene expressions, binding targets and binding motifs of large number of RNA binding proteins (RBPs) with a co-methylation network constructed using large-scale m6A methylomes across diverse cell states. We applied the framework and successfully identified 32 high-confidence m6A regulators that modulated the variable m6A sites away from stop codons in a cell-specific manner. To validate them, we knocked down three regulators respectively and found two of them (TRA2A and CAPRIN1) selectively promoted the methylations of the m6A sites co-localized with their binding targets on RNAs through physical interactions with the m6A writers. Knockdown of TRA2A increased the stabilities of the RNAs with TRA2A bound near the m6A sites and decreased the viability of cells. The successful identification of m6A regulators demonstrates a powerful and widely applicable strategy to elucidate the cell-specific m6A regulators. Additionally, our discovery of pervasive trans-acting regulating of m6A provides novel insights into the mechanisms by which spatial and temporal dynamics of m6A methylomes are established.

DNA methylation is one of the most important epigenetic mechanisms to regulate gene expression, which is highly dynamic during development and specifically maintained in somatic cells. Aberrant DNA methylation patterns are strongly associated with human diseases including cancer. How are the cell-specific DNA methylation patterns established or disturbed is a pivotal question in developmental biology and cancer epigenetics. Currently, compelling evidence has emerged that long non-coding RNA (lncRNA) mediates DNA methylation in both physiological and pathological conditions. In this review, we provide an overview of the current understanding of lncRNA-mediated DNA methylation, with emphasis on the roles of this mechanism in cancer, which to the best of our knowledge, has not been systematically summarized. In addition, we also discuss the potential clinical applications of this mechanism in RNA-targeting drug development.

Herein, we report the discovery and characterization of a novel class of PAK4 inhibitors with a quinazoline scaffold. Based on the shape and chemical composition of the ATP-binding pocket of PAKs, we chose a 2,4-diaminoquinazoline series of inhibitors as a starting point. Guided by X-ray crystallography and a structure-based drug design (SBDD) approach, a series of novel 4-aminoquinazoline-2-carboxamide PAK4 inhibitors were designed and synthesized. The inhibitors' selectivity, therapeutic potency, and pharmaceutical properties were optimized. One of the best compounds, 31 (CZh226), showed remarkable PAK4 selectivity (346-fold vs PAK1) and favorable kinase selectivity profile. Moreover, this compound potently inhibited the migration and invasion of A549 tumor cells by regulating the PAK4-directed downstream signaling pathways in vitro. Taken together, these data support the further development of 31 as a lead compound for PAK4-targeted anticancer drug discovery and as a valuable research probe for the further biological investigation of group II PAKs.

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Tandem repeats (TRs) are abundant and widely distributed in eukaryotic genomes. TRs are thought to have various functions in gene transcription, DNA methylation, nucleosome position and chromatin organization. Variation of repeat units in the genome is observed in association with a number of diseases, such as Fragile X Syndrome, Huntington's disease and Friedreich's ataxia. However, the underlying mechanisms involved are poorly understood, largely owing to the technical limitations in modification of TRs at definite sites in the genome in vivo. Transcription activator-like effector nucleases (TALENs) are widely used in recent years in gene targeting for their specific binding to target sequences when engineered in vitro. Here, we show that the repair of a double-strand break (DSB) induced by TALENs adjacent to a TR can produce serial types of mutations in the TR region. Sequencing analysis revealed that there are three types of mutations induced by the DSB repair, including indels only within the TR region or within the flanking TALEN target region or simutaneously within both regions. Therefore, desired TR mutant types can be conveniently obtained by using engineered TALENs. These results demonstrate that TALENs can serve as a convenient tool for modifying TRs in the genome in studying the functions of TRs.

A new series of novel 1-phenanthryl-tetrahydroisoquinoline derivatives were designed, synthesized and biologically evaluated for their PAK4 inhibitory activities and anti-proliferative effects against three cancer cell lines A549, MCF-7 and HT-1080. Among them, compound 12a exhibited the most potent inhibitory activity against PAK4 with an IC50 value of 0.42 μM. Moreover, this compound inhibited the invasion of A549 tumor cells by regulating the PAK4-LIMK1-cofilin signaling pathway in vitro, and exhibited anti-tumor activity in vivo in the A549 tumor xenograft model. To further evaluate the binding mode of 12a with PAK4, the biotinylated 12a derivative has been synthesized and it was used for immunoprecipitation assay. Intriguingly, our observations suggest that 12a interacts with both the N- and C-termini of PAK4.