Analysis of rpoS and bolA gene expression under various stress-induced environments in planktonic and biofilm phase using 2−ΔΔCT method

Abstract

Genetic adaptation is one of the key features of Escherichia coli (E. coli) that ensure its survival in different hostile environments. E. coli seems to initiate biofilm development in response to specific environmental cues. A number of properties inherent within bacterial biofilms indicate that their gene expression is different from that of planktonic bacteria. Two of the possible important genes are rpoS and bolA. The rpoS gene has been known as the alternative sigma (σ) factor, which controls the expression of a large number of genes, which are involved in responses to a varied number of stresses, as well as transition to stationary phase from exponential form of growth. Morphogene bolA response to stress environment leads to round morphology of E. coli cells, but little is known about its involvement in biofilms and its development or maintenance. The purpose of this study was to understand and analyse the responses of rpoS and bolA gene to sudden change in the environment. In this study, E. coli K-12 MG1655, rpoS, and bolA mutant strains were used and gene expression was studied. Results show that both genes contribute to the ability to respond and adapt in response to various types of stresses. RpoS response to various stress environments was somehow constant in both the planktonic and biofilm phases, whereas bolA responded well under various stress conditions, in both planktonic and biofilm mode, up to 5-6-fold change in the expression was noticed in the case of pH variation and hydrogen peroxide stress (H(2)O(2)) as compared with rpoS.

Enhanced cadmium resistance and accumulation inPseudomonas putidaKT2440 expressing the phytochelatin synthase gene ofSchizosaccharomyces pombe

“…Expression level was characterized by relative real‐time PCR using method (Adnan et al . ). Housekeeping gene 16S rRNA gene was used as the internal control.…”

Genome Evolution in the Cold: Antarctic Icefish Muscle Transcriptome Reveals Selective Duplications Increasing Mitochondrial Function

“…Other noteworthy duplicated genes are: 1) bolA homolog 1 ( Escherichia coli ) ( bola1 ), which is well studied in E. coli and is involved in the response and adaptation to various stress conditions, among which are pH variation and oxidative stress (Adnan et al 2011); 2) cystatin A ( csta ), encoding a cysteine proteinase that restrains UVB-induced apoptosis of keratinocytes and confers enhanced resistance to high salt concentrations, drought, oxidative, and cold stresses in plants (Takahashi et al 2007; Zhang et al 2008); 3) nuclear protein 1 ( nupr1 ), encoding a key player in cellular stress response (Goruppi and Iovanna 2010), which is upregulated in the liver of rainbow trout during stress response and recovery (Momoda et al 2007); 4) ring finger protein 7 ( rnf7 ), encoding a redox-inducible and apoptosis-protective antioxidant protein, which also decreases endogenous ROS production and oxidative DNA damage after exposure to heat shock when overexpressed (Lee et al 2008; Sun 2008); 5) defender against cell death 1 ( dad1 ), which codes for a subunit of the oligosaccharyltransferase complex and is upregulated in zebrafish embryonic cell line by XBP-1S, a key transcriptional regulator of the unfolded protein response that allows the recovery of endoplasmic reticulum (ER) function (Hu et al 2007); and 6) chaC, cation transport regulator-like 1 ( chac1 ), which encodes a component of the mammalian unfolded protein response pathway (Mungrue et al 2009). Duplicated genes were found for four additional proteins involved in protein folding: Prefoldin subunit 2 (Pfdn2), which is a subunit of the molecular chaperone prefoldin involved in the folding of newly synthesized proteins, mainly actins and tubulins (Pfdn2 subunit is thought to facilitate the formation of the prefoldin complex and also contain a DNA binding motif) (Martin-Benito et al 2002; Miyazawa et al 2011); Thioredoxin domain containing 9 (Txndc9), which, curiously, has an action antagonistic to that of prefoldin and the combined action of the two proteins is thought to be crucial to establishing a functional cytoskeleton (Stirling et al 2006); Ubiquitin fusion degradation 1-like (Ufd1l), which is part of a multiprotein complex involved in the recognition and export of polyubiquitin-tagged misfolded proteins from the ER to the cytoplasm, where they are degraded by the 26S proteasome (Bays and Hampton 2002); Ufd1l subunit is the one that directly interacts and stimulates the activity of ubiquitin ligase gp78, which mediates the degradation of misfolded ER proteins (Cao et al 2007) and is also induced in yeast in response to cold (Schade et al 2004); ubiquitin-like 5 ( ubl5 ), which is upregulated in response to mitochondrial stress, is required for mounting mitochondrial unfolded protein response, leading to the induction and subsequent import into mitochondria of mitochondrial chaperones (Haynes and Ron 2010).…”

Elucidating the role of highly homologous Nicotiana benthamiana ubiquitin E2 gene family members in plant immunity through an improved virus-induced gene silencing approach

“…qRT-PCR was performed using gene specific primers and SYBR Green (Life Technologies) on the LightCycler ® 480 Instrument II (Roche), with the NbEF1α gene being used as an internal reference. The relative expression level of NbRbohB gene was calculated using the 2 −ΔΔ CT  method as described [65]. The primers used for qRT-PCR are shown in the Additional file 2: Table 1.…”