Integrative single-cell and bulk transcriptomic analyses identify DRAM1 as a candidate gene from fibroblast-associated transcriptional programs in colorectal cancer.

Colorectal cancer (CRC) is characterized by marked cellular heterogeneity and dynamic remodeling of the tumor microenvironment. Fibroblasts represent an important stromal component of CRC and may contribute to tumor progression through complex transcriptional programs and intercellular interactions. However, fibroblast-associated candidate genes with potential biological relevance remain incompletely defined. We applied an integrative strategy combining publicly available single-cell RNA sequencing (scRNA-seq), pseudotime analysis, CellChat, high-dimensional weighted gene co-expression network analysis (hdWGCNA), bulk transcriptomic validation, machine learning prioritization, and experimental assays. Using the GSE221575 dataset, we mapped the cellular landscape of CRC and adjacent normal tissues and identified fibroblast-associated co-expression modules. Candidate genes were further prioritized across five independent GEO bulk transcriptomic cohorts. DRAM1 was selected for validation by qRT-PCR, western blotting, proliferation, colony formation, wound-healing, Transwell, and xenograft assays. scRNA-seq analysis identified 17 cell clusters corresponding to 10 major cell lineages in CRC and adjacent normal tissues. Cell-cell communication analysis showed extensive signaling interactions among stromal, epithelial, endothelial, and immune cell populations, with fibroblasts occupying a prominent position in the communication network. hdWGCNA identified 31 fibroblast-associated modules, from which 123 candidate genes were extracted. Integrative bulk validation and machine learning analysis yielded five core genes, including INHBA, COL6A3, SPARC, DRAM1, and COL1A2. Among them, DRAM1 was selected for experimental validation. DRAM1 expression was elevated in CRC tissues and selected CRC cell lines. Loss-of-function assays showed that DRAM1 silencing enhanced CRC cell proliferation, migration, invasion, and xenograft growth, suggesting that increased DRAM1 expression may reflect a compensatory stress-response program rather than a purely oncogenic function. This study provides an integrative characterization of fibroblast-associated transcriptional programs in CRC and identifies DRAM1 as a candidate gene emerging from these programs. Functional assays support a tumor-restraining role for DRAM1 in CRC epithelial tumor cell models, whereas its direct role in fibroblast-mediated stromal-immune regulation requires further investigation.