In Arabidopsis, secondary cell walls (SCW) are formed in fiber cells and vessel cells in vascular tissue for providing plants with mechanical strength and channels for the long distance transportation of water and nutrients. NST1 acts as a key gene for the initiation of SCW formation through a hierarchy transcriptional network. In this study, we report that NST activity is modulated by XND1 during plant growth. Using yeast two hybrid screening and in vivo protein interaction analysis, XND1 was identified as an NST1-interacting protein to modulate NST1 activity. XND1 and NST1 were co-localized in the nucleus and the interaction of XND1 with NST1 resulted in inhibition of NST1 transactivation activity. In the process of inflorescence growth, XND1 was expressed in a similar pattern with NST1. Up-regulation of XND1 in fiber cells repressed their SCW formation. The study demonstrates that NST1 activity is modulated by XND1 in regulation of SCW formation.