Abstract This study applied synchrotron radiation XRD to analyze the mechanical behavior of cellulose microfibrils in wood containing annual rings (thickness: 5 mm), for different layers of the secondary cell wall, under uniaxial load. Cellulose in S2 and in S1 and S3 layers were analyzed respectively, and the data were used to investigate for deformation behavior in the lattice spacing (d 004). As a result, the mechanical behavior of cellulose sometimes differed from the behavior of bulk wood. The rigidity of cellulose in the S2 layer was larger than in S1 and S3 layers under both of tensile and compressive loads. However, once standardized with respect to estimated cellulose amount, this standardized rigidity was comparable across all layers and loading conditions. Variation in microfibril angle (MFA) and lattice spacing (d 004) of cellulose barely changed at all under compressive load. Under tensile loads, there were both of positive and negative changes in MFA variation in both S2 layer and S1 and S3 layers, while d 004 variation had little changes in almost all cases.