To determine the effect of duration of dietary vitamin A restriction on site of fat deposition in growing cattle, 60 Holstein steers (BW = 218.4 +/- 6.55 kg) were fed a diet based on high-moisture corn, with 2,200 IU of supplemental vitamin A/kg of DM (control) or no supplemental vitamin A for a long (243 d; LR) or short (131 d; SR) restriction before slaughter at 243 d. The SR steers were fed the control diet for the first 112 d. Steers were penned individually and fed for ad libitum intake. Jugular vein blood samples for serum retinol analysis were collected on d 1, 112, and 243. Carcass samples were collected for composition analysis. Subcutaneous fat samples were collected for fatty acid composition. Fat samples from the i.m. and s.c. depots were collected to measure adipocyte size and density. Feedlot performance (ADG, DMI, and G:F) was not affected (P > 0.05) by vitamin A restriction. On d 243, the i.m. fat content of the LM was 33% greater (P < 0.05) for LR than for SR and control steers (5.6 vs. 3.9 and 4.2% ether extract, respectively). Depth of back-fat and KPH percentage were not affected (P = 0.44 and 0.80, respectively) by vitamin A restriction. Carcass weight, composition of edible carcass, and yield grade were similar among treatments (P > 0.10). Liver retinol (LR = 6.1, SR = 6.5, and control = 44.7 microg/g; P < 0.01) was reduced in LR and SR vs. control steers. On d 243, LR and SR steers had similar serum retinol concentrations, and these were lower (P < 0.01) than those of control steers (LR = 21.2, SR = 25.2, and control = 36.9 microg/dL). Intramuscular adipose cellularity (adipocytes/mm2 and mean adipocyte diameter) on d 112 and 243 was not affected (P > 0.10) by vitamin A restriction. Restricting vitamin A intake for 243 d increased i.m. fat percentage without affecting s.c. or visceral fat deposition, feedlot performance, or carcass weight. Restricting vitamin A intake for 131 d at the end of the finishing period appears to be insufficient to affect the site of fat deposition in Holstein steers.