Fig. 4

Stress mechanisms and cellular processes involved in the occurrence of stress symptoms when (over)expressing (heterologous) (membrane) proteins. (1) Decreased growth rate: (a) ppGpp destabilises \(\sigma ^{70}\) promoter complexes and frees RNAP; (b) Increased amount of alternative sigma factors in the cell; (c) Increased transcription of genes dependent on alternative sigma factors; (d) ppGpp interferes in all processes of ribosome assembly and translation; (e) Less translation of metabolic proteins; (f) Translation of proteins of interest sequester part of the ribosomes; (g) Amino acid starvation induces differential charging of tRNAs, benefitting optimal codons; (h) Cpx response reduces energy generation which is negative for growth; (i) Phage shock response induces the production of acetate negatively impacting growth; (2) Impaired protein synthesis: (a) Increased translation errors lead to misfolded proteins; (b) Codon optimised sequences negatively impact folding of proteins and thus increase misfolded proteins; (c) Changing the mRNA sequence of genes changes their stability, which impacts translation; (d) Amino acid starvation induces differential charging of tRNAs, benefitting optimal codons; (e) The lack of rare codons negatively impacts expression of heterologous genes; (f) ppGpp together with increased amounts of alternative sigma factors decreases transcription from \(\sigma ^{70}\) promoters and away from the gene of interest; (g) ppGpp inhibits GTPase activity, having a negative influence on insertion of proteins into the membrane; (h) Overloading of the Sec translocon has increased amounts of IB and misfolded proteins as a consequence; (i) Aggregation of elongation factor Tu, reduces the delivery of tRNAs to the ribosome. (3) Genetic instability: (a) Non-mutagenic DSB repair in unstressed cells; (b) Spontaneous DSB in growing E. coli; (c) \(\sigma ^{\textrm{E}}\) increases the occurrence of DSB; (d) ppGpp inhibits DNA replication; (e) DSB and stalled DNA replication induce the SOS response; (f) Error-prone DNAPs are synthesised; (g) DSB induce iraD transcription and IraD stabilises \(\sigma ^{\textrm{S}}\); (h) \(\sigma ^{\textrm{S}}\) promotes the use of error-prone DNAP in DSB repair, increasing the mutation rate. (4) Aberrant cell size: (a) Filamentation of cells; (b) FtsZ locates mid-cell; (c) MinCDE assists FtsZ in localising mid-cell; (d) ppGpp inhibits GTPase activity of FtsZ; (e) SOS response activates cell division inhibitors SulA and YmfM; (f) MinD aggregates in IB, impacting cell division; (g) The Cpx response and the phage shock response negatively influence energy generation and high energy processes respectively, reducing cell division processes. \(\sigma\) sigma factor, ppGpp guanosine tetra- and pentaphosphate, RNAP RNA polymerase, IB inclusion bodies, rRNA ribosomal RNA, mRNA messenger RNA, tRNA transfer RNA, DNAP DNA polymerase, DSB double stranded DNA break. Gene structures are displayed according to the SBOL guidelines [6]