DataSheet1_Gene Co-Expression Network Analysis Identifies Vitamin D-Associated Gene Modules in Adult Normal Rectal Epithelium Following Supplementation.xlsx

Colorectal cancer (CRC) is a common, multifactorial disease. While observational studies have identified an association between lower vitamin D and higher CRC risk, supplementation trials have been inconclusive and the mechanisms by which vitamin D may modulate CRC risk are not well understood. We sought to perform a weighted gene co-expression network analysis (WGCNA) to identify modules present after vitamin D supplementation (when plasma vitamin D level was sufficient) which were absent before supplementation, and then to identify influential genes in those modules. The transcriptome from normal rectal mucosa biopsies of 49 individuals free from CRC were assessed before and after 12 weeks of 3200IU/day vitamin D (Fultium-D3) supplementation using paired-end total RNAseq. While the effects on expression patterns following vitamin D supplementation were subtle, WGCNA identified highly correlated genes forming gene modules. Four of the 17 modules identified in the post-vitamin D network were not preserved in the pre-vitamin D network, shedding new light on the biochemical impact of supplementation. These modules were enriched for GO terms related to the immune system, hormone metabolism, cell growth and RNA metabolism. Across the four treatment-associated modules, 51 hub genes were identified, with enrichment of 40 different transcription factor motifs in promoter regions of those genes, including VDR:RXR. Six of the hub genes were nominally differentially expressed in studies of vitamin D effects on adult normal mucosa organoids: LCN2, HLA-C, AIF1L, PTPRU, PDE4B and IFI6. By taking a gene-correlation network approach, we have described vitamin D induced changes to gene modules in normal human rectal epithelium in vivo, the target tissue from which CRC develops.

结直肠癌（CRC）是一种常见且多因素引起的疾病。尽管观察性研究已确定维生素D水平降低与CRC风险增加之间存在关联，但补充剂试验的结果尚无定论，而维生素D可能调节CRC风险的作用机制尚不明确。本研究旨在通过加权基因共表达网络分析（WGCNA）来识别在维生素D补充后（血浆维生素D水平充足时）存在的而在补充前不存在的模块，然后识别这些模块中的关键基因。对49名无CRC个体正常直肠黏膜活检的转录组进行了评估，这些个体在补充3200IU/天维生素D（Fultium-D3）前后的12周内使用双末端总RNA测序。尽管维生素D补充后对表达模式的影响微乎其微，但WGCNA识别出高度相关的基因形成基因模块。在维生素D补充后的网络中识别的17个模块中，有4个在维生素D补充前的网络中未保留，从而为补充剂的生化影响提供了新的见解。这些模块富含与免疫系统、激素代谢、细胞生长和RNA代谢相关的GO术语。在四个与治疗相关的模块中，共确定了51个枢纽基因，这些基因启动子区域中富集了40种不同的转录因子基序，包括VDR:RXR。在研究维生素D对成年正常黏膜类器官的影响的研究中，有6个枢纽基因在名义上存在差异表达：LCN2、HLA-C、AIF1L、PTPRU、PDE4B和IFI6。通过采用基因相关性网络方法，我们描述了维生素D在正常人类直肠上皮中的诱导基因模块变化，这是CRC发展的目标组织。