Enhanced protein secretion in reduced genome strains of Streptomyces lividans

S. lividans TK24 is a popular host for the production of small molecules and for the secretion of heterologous proteins. TK24 has a large genome with at least 29 secondary metabolite gene clusters that are non-essential for viability and undergo complex regulation. To optimize heterologous protein secretion, we previously constructed ten chassis strains that are devoid of several secondary metabolite gene clusters. Genome reduction was aimed at reducing carbon flow to secondary metabolites and pigmentation in the spent growth medium and improving colony morphology. Strains RG1.0-RG1.10 contain various deletion combinations of the blue actinorhodin cluster (act), the calcium-dependent antibiotic (cda), the undecylprodigiosin (red) and coelimycin A (cpk) clusters, the melanin cluster (mel), the matAB genes that affect mycelial aggregation and the non-essential sigma factor hrdD that controls the transcription of Act and Red regulatory proteins. Two derivative strains, RG1.5 and 1.9, showed a [~]15% reduction in growth rate, >2-fold increase in the total mass yield of their native secretome and altered abundance of several specific proteins compared with TK24. Metabolomics and RNAseq analysis revealed that genome reduction led to rapid cessation of growth due to aminoacid depletion and caused both redox and cell envelope stresses, upregulation of the Sec-pathway components secDF and chaperones and a cell envelope two component regulator. RG1.9 maintained elevated heterologous secretion of mRFP and mTNF by 12-70%. An integrated model is presented linking genome reduction and enhanced secretion.