Drought stress triggers remarkable physiological changes and growth impediments, which significantly diminish plant biomass and crop yield. However, certain plant species show notable resilience, maintaining nearly normal yields under severe water deficits. For example, sorghum is a naturally drought-tolerant crop, which is ideal for studying plant adaptive responses to drought. Here we used sorbitol treatments to simulate drought-induced osmotic stress in sorghum cell suspension cultures and analysed fractions enriched for extracellular matrix proteins using isobaric tags for relative and absolute quantification technology. Sorbitol induced an overall increase in protein secretion, with putative redox proteins, proteases, and glycosyl hydrolases featuring prominently among the responsive proteins. Gene expression analysis of selected candidates revealed regulation at the transcriptional level. There was a notable differential gene expression between drought-tolerant and drought-sensitive sorghum varieties for some of the candidates. This study shows that protein secretion is a major component of the sorghum response to osmotic stress. Additionally, our data provide candidate genes, which may have putative functions in sorghum drought tolerance, and offer a pool of genes that could be developed as potential biomarkers for rapid identification of drought tolerant lines in plant breeding programs.