To understand better how wheat chloroplasts respond to salt stress, we conducted a comparative proteomics analysis. Seedlings (12-day-old) were exposed to 75 mM NaCl for 1, 2, or 3 d. Accumulation of Na+ ions were rapid and excessive. Decreases were recorded for photosynthesis and transpiration rates, stomatal conductance, and relative water content whereas the level of proline was increased. Protein abundance within the chloroplasts was examined by two-dimensional electrophoresis. More than 100 protein spots reproducibly detected on each gel, 21 differential spots were expressed during salt treatment. Using LTQ-FTICR hybrid mass spectrometry, we assigned identities to 65 unique proteins for those differentially abundant spots. Most proteins were up-regulated at 2 and 3 d after being down-regulated at Day 1. Others showed only slight responses after 3 d of treatment, including those for Rubisco, glutamate dehydrogenase, glyceraldehyde-3-phosphate dehydrogenase, isocitrate dehydrogenase, photosystem I, and pyridoxal biosynthesis. The abundance of the ATP synthase (alpha, Beta, and Epsilon) and V-type proton ATPase subunits down-regulated, and impersonated the mechanism, resulting in a negative impact by Na+ on the photosynthetic machinery. This temporary increase and subsequent decrease in protein contents may demonstrate a counterbalancing influence of identified proteins. Several proteins such as Cyt b6f, germin-like-protein, glutathione peroxidase, ATP synthase subunit gamma, glutamine synthetase, fructose-bisphosphate aldolase, S-adenosylmethionine synthase, carbonic anhydrase were gradually up-regulated during our treatment period, which is identified as marker proteins.