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We investigate if and how diets of gray wolves from the Yukon Territory, Canada, have changed from the Pleistocene (>52.8 ka BP to 26.5 ka BP [±170 y BP]) to the recent Holocene (1960s) using dental microwear analysis of carnassial teeth and stable isotope analyses of carbonates (δ13CCO3 and δ18OCO3) and collagen (δ13Ccol and δ15Ncol) from bone. We find that dental microwear patterns are similar between the Pleistocene and Holocene specimens, indicating that there has been no change in carcass utilization behaviours, where flesh, not bone, is primarily consumed. Based on minimal changes in δ13CCO3 and δ13Ccol values, we find that, over thousands of years, Yukon gray wolves have remained generalist predators feeding upon several large ungulate species. Interestingly, δ15Ncol values suggest that the extinction of megafaunal species at ~11.7 Ka induced a shift from a diet comprised primarily of horse (Equus sp.) to one based on cervids (i.e. moose and caribou). Survival of large-bodied cervids, such as caribou (Rangifer tarandus), was likely key to wolf survival. Although gray wolves survived the end Pleistocene megafauna extinction and demonstrate a degree of ecological flexibility, we suggest that failure to preserve major elements of their current niche (e.G. caribou) may result in continued population declines, especially in the face of increasing anthropogenic influences.