Extracellular matrix (ECM) is the first line of defense which is an inimitable organelle that perceives biotic and abiotic stresses and reprograms biological processes of host. It also activates innate immune responses in temporal and spatial manner and acts as physical scaffold that prevents the entry of pathogen and microbes in response to stress and hormonal signals. Stem rot, caused by Macrophomina phaseolina adversely affects fiber production in jute. However, how wall related susceptibility affects ECM proteome remains undetermined in bast fiber crops. Here, stem rot responsive quantitative temporal ECM proteome was developed in jute upon M. phaseolina infection. Using isobaric tags for relative and absolute quantitative proteomics and MS/MS analysis, we identified 415 disease responsive proteins (DRPs), involved in wall integrity, acidification, proteostasis, hydration and redox homeostasis. Disease-related correlation network identified functional hubs related to wall degrading enzymes, structural carbohydrates and signaling govern rot responsive wall-susceptibility.