Dietary methionine restriction impairs anti-tumor immunity through gut microbiota

Dietary methionine restriction represses growth and improves therapeutic responses in several pre-clinical settings. However, how this dietary intervention impacts cancer progression in the context of the immune system is unknown. Here we analyzed the CD45+ immune cells from the small intestine of control (CTRL) diet or methionine-restricted (MR) diet fed tumor-free C57BL/6J donor mice and tumor-bearing Apc recipient mice transplanated with feces from these diet-fed tumor-free C57BL/6J mice by scRNA-seq. Our analysis indicate that fecal microbes from methionine-restricted tumor-free C57BL/6J mice are sufficient to represss T cell activation in the small intestine of Apc mice. Overall design: CD45+ immune cells purified from the small intestine of diet-fed C57BL6/J donor mice and fecal transplanted Apc mice, with 2 mice per sample, were counted and examined for viability with trypan blue staining using a TC-20 cell counter (Bio-Rad). Approximately 10,000 live cells with above 70% viability were loaded into the Single Cell Chip to generate single cell emulsion in Chromium Controller (10x Genomics, 120263) with Chromium Single Cell 3' Library & Gel Bead Kit v3.1 (10x Genomics, 1000268). Reverse transcription of mRNA and cDNA amplification were carried out following the manufacture's instruction (10x Genomics, 1000268). The amplified cDNA was further fragmented to construct NGS libraries. The libraries were then sequenced by the NIEHS Epigenomics and DNA Sequencing Core Laboratory with the parameters recommended in the manufacture's instruction.