Microbial bioremediation of Cr(VI)-contaminated environments has drawn extensive concern. However, the molecular processes underlying the microbial Cr(VI) tolerance and reduction remain unclear. We isolated a Cr(VI)-reducing Lysinibacillus fusiformis strain 15-4 from soil on the Qinghai-Tibet Plateau. When grown in 1 mM and 2 mM Cr(VI)-containing medium, strain 15-4 could reduce 100% and 93.7% of Cr(VI) to Cr(III) after 36 h and 60 h of incubation, respectively. To know the molecular processes in response to Cr(VI), transcriptome sequencing was carried out using RNA-Seq technology. The results annotated a total of 3913 expressed genes in the strain. One thousand ninety-eight genes (28.1%) were significantly (fold change >= 2, false discovery rate <= 0.05) expressed in response to Cr(VI), of which 605 (55.1%) were upregulated and 493 (44.9%) were downregulated. The enrichment analysis showed that a total of 630 differentially expressed genes (DEGs) were enriched to 122 KEGG pathways, of which 8 pathways were significantly (p < 0.05) enriched in Cr(VI)-treated sample, including ATP-binding cassette (ABC) transporters (97 DEGs), ribosome (40), sulfur metabolism (16), aminoacyl-tRNA biosynthesis (19), porphyrin metabolism (20), quorum sensing (44), oxidative phosphorylation (17), and histidine metabolism (10), suggesting that these pathways play key roles to cope with Cr(VI) in the strain. The highly upregulated DEGs consisted of 29 oxidoreductase, 18 dehydrogenase, 14 cell redox homeostasis and stress response protein, and 10 DNA damage and repair protein genes. However, seven Na+:H+ antiporter complex-coding DEGs and most of transcriptional regulator-coding DEGs were significantly downregulated in the Cr-treated sample. Many of FMN/NAD(P)H-dependent reductase-encoding genes were greatly induced by Cr, suggesting the involvement of these genes in Cr(VI) reduction in strain 15-4. Sulfur and iron ions as well as the thiol-disulfide exchange reactions might play synergistic roles in Cr reduction.