5-Methylcytosine (mC) is a well-characterized DNA modification, and is also predominantly reported in abundant non-coding RNAs in both prokaryotes and eukaryotes. However, the distribution and biological functions of mC in plant mRNAs remain largely unknown. Here, we report transcriptome-wide profiling of RNA mC in Arabidopsis thaliana by applying mC RNA immunoprecipitation followed by a deep-sequencing approach (mC-RIP-seq). LC-MS/MS and dot blot analyses reveal a dynamic pattern of mC mRNA modification in various tissues and at different developmental stages. mC-RIP-seq analysis identified 6045 mC peaks in 4465 expressed genes in young seedlings. We found that mC is enriched in coding sequences with two peaks located immediately after start codons and before stop codons, and is associated with mRNAs with low translation activity. We further demonstrated that an RNA (cytosine-5)-methyltransferase, tRNA-specific methyltransferase 4B (TRM4B), exhibits mC RNA methyltransferase activity. Mutations in TRM4B display defects in root development and decreased mC peaks. TRM4B affects the transcript levels of the genes involved in root development, which is positively correlated with their mRNA stability and mC levels. Our results suggest that mC in mRNA is a new epitranscriptome marker inArabidopsis, and that regulation of this modification is an integral part of gene regulatory networks underlying plant development.

Wound healing is a dynamic process involving gene-expression changes that drive re-epithelialization. Here, we describe an essential role for polyamine regulator AMD1 in driving cell migration at the wound edge. The polyamines, putrescine, spermidine, and spermine are small cationic molecules that play essential roles in many cellular processes. We demonstrate that AMD1 is rapidly upregulated following wounding in human skin biopsies. Knockdown of AMD1 with small hairpin RNAs causes a delay in cell migration that is rescued by the addition of spermine. We further show that spermine can promote cell migration in keratinocytes and in human ex vivo wounds, where it significantly increases epithelial tongue migration. Knockdown of AMD1 prevents the upregulation of urokinase-type plasminogen activator/urokinase-type plasminogen activator receptor on wounding and results in a failure in actin cytoskeletal reorganization at the wound edge. We demonstrate that keratinocytes respond to wounding by modulating polyamine regulator AMD1 in order to regulate downstream gene expression and promote cell migration. This article highlights a previously unreported role for the regulation of polyamine levels and ratios in cellular behavior and fate.

Caspase recuitment domain-containing protein 9 (CARD9) functions in different inflammation pathways to elicit responses to microbial signals and is known to affect intestinal inflammation. Examining the APC min mouse model of intestinal tumorigenesis and using stringently controlled, sex-and age-matched pairs of CARD9-competent and CARD9-deficient mice, we have found that CARD9 has a restricted but strong effect on tumorigenesis in the large intestine. We have found that CARD9 reduces viability specifically in males and promotes tumorigenesis specifically in the large intestines of these male mice. To our knowledge, this is the first gene ablation in APC min mice that solely affects colon tumors in male subjects and, as such, may have significant clinical implications. Additional data suggest correlative disruption of plasma cytokine expression and immune infiltration of the tumors. We speculate that known sex-specific differences in human colorectal cancer may involve inflammation, particularly CARD9-dependent inflammation. Cancer Immunol Res; 3(7); 721-6. Ó2015 AACR.