# The Production of Curli Amyloid Fibers Is Deeply Integrated into the Biology of _Escherichia coli_
## 2 Results and Discussion
### CsgD Transcript and Protein Levels Are Altered in Several Strains with CR Phenotypes
It is also worth noting that a threefold increase in _csgD_ transcripts was observed in _cysB_, but CsgD protein levels were half of WT, whereas _fes_ was observed to have _csgD_ transcript levels nearly identical to wild-type with approximately half the levels of CsgD protein (Table S9). The _csgD_ transcript data presented here agree with some previously published findings. For instance, mutants in pyrimidine synthesis, _pyrC_, were shown to have decreased _csgD_ transcripts [131] and we observed similar decreases in _csgD_ expression in a _purD_ mutant (Table 2). It is interesting to speculate that these two mutants act through the same mechanism, but more work is needed to elucidate how _pyrC_ and _purD_ contribute to curli biogenesis. Hfq, a chaperone protein responsible for RNA stability, is required for _csgD_ expression in _S. enterica_[132], and we confirmed that this is true in _E. coli_ (Table 2). Additionally, RcsB represses the _fhlDC_ operon [133], induces expression of \begin{table}
\begin{tabular}{c c c c c c} \hline \hline
**Strain** & **Average _csgD_ Levels & **Std Dev** & _p-Value_ & **Average 16s Levels** & **Std Dev** & _p-Value_ \\ \hline BW25113 & \(3.141\times 10^{-2}\) & \(1.652\times 10^{-2}\) & N/A & \(1.38\times 10^{-2}\) & \(7.56\times 10^{-3}\) & N/A \\ _dam_ & \(4.605\times 10^{-2}\) & \(1.173\times 10^{-2}\) & \(0.107\) & \(2.05\times 10^{-2}\) & \(4.31\times 10^{-3}\) & \(8.93\times 10^{-2}\) \\ _fftq_ & \(1.742\times 10^{-4}\) & \(3.279\times 10^{-5}\) & \(9.36\times 10^{-4}\) & \(6.46\times 10^{-3}\) & \(6.48\times 10^{-3}\) & \(1.02\times 10^{-1}\) \\ _nagA_ & \(2.407\times 10^{-2}\) & \(2.143\times 10^{-3}\) & \(0.306\) & \(4.79\times 10^{-3}\) & \(4.23\times 10^{-3}\) & \(2.94\times 10^{-2}\) \\ _dksA_ & \(3.777\times 10^{-2}\) & \(9.779\times 10^{-3}\) & \(0.436\) & \(3.14\times 10^{-3}\) & \(8.50\times 10^{-4}\) & \(6.51\times 10^{-3}\) \\ \hline \hline \end{tabular} Std Dev: Standard deviation.  
\end{table} 
Table 4: _csgD_ levels altered by decreased ribosome RNA levels. \begin{table}
\begin{tabular}{c c c c c} \hline \hline
**CR Phenotype** & **Strain** & **Average Relative to BW25113 (WT)** & **Standard Deviation** & **Outcome of _t-Test_** \\ \hline Light Red & _priA_ & 0.023 & \(\pm 0.004\) & _p-value_ \(<0.001\) \\ Light Red & _aneR_ & 2.324 & \(\pm 0.755\) & _p-value_ \(<0.05\) \\ Light Red & _glycG_ & 1.316 & \(\pm 0.175\) & Not significant \\ Light Red & _cmr_ & 1.586 & \(\pm 0.561\) & Not significant \\ Light Red & _dam_ & 0.967 & \(\pm 0.268\) & Not significant \\ Light Red & _hdfR_ & 0.752 & \(\pm 0.254\) & Not significant \\ Light Red & _mltA_ & 0.874 & \(\pm 0.170\) & Not significant \\ Dark Red & _cysB_ & 3.06 & \(\pm 0.982\) & _p-value_ \(<0.001\) \\ Dark Red & _pcnB_ & 1.602 & \(\pm 0.354\) & _p-value_ \(<0.05\) \\ Dark Red & _trnB_ & 1.371 & \(\pm 0.240\) & Not significant \\ Dark Red & _rcsB_ & 3.723 & \(\pm 0.970\) & _p-value_ \(<0.05\) \\ Dark Red & _sdiA_ & 2.952 & \(\pm 0.684\) & _p-value_ \(<0.05\) \\ Dark Red & _fes_ & 0.999 & \(\pm 0.217\) & Not significant \\ Dark Red & _muA_ & 2.302 & \(\pm 0.678\) & Not significant \\ Dark Red & _qscC_ & 2.861 & \(\pm 2.190\) & Not significant \\ Dark Red & _arcA_ & 1.603 & \(\pm 0.219\) & _p-value_ \(<0.05\) \\ Dark Red & _mdoC_ & 1.581 & \(\pm 0.309\) & _p-value_ \(<0.05\) \\ Dark Red & _perR_ & 1.923 & \(\pm 0.537\) & _p-value_ \(<0.05\) \\ Dark Red & _cusB_ & 2.224 & \(\pm 0.998\) & _p-value_ \(<0.05\) \\ \hline \hline \end{tabular}  
* * Increased _csgD_ transcript levels driven by decrease in 16s ribosomal RNA (rRNA) levels (see Table 4). CR denotes Congo red (CR) binding phenotype.  
\end{table} 
Table 2: _Cont._ \begin{table}
\begin{tabular}{c c} \hline \hline
**Outcome of _t-Test** & **Number of Strains** \\ \hline Not significant & 15 \\ _p-_value \(<0.05\) & 14 \\ _p-_value \(<0.001\) & 7 \\ Uncertain & 2 \\ Total & 38 \\ \hline \hline \end{tabular}
\end{table} 
Table 3: _csgD_ qualitative PCR significance for selected mutants. the small RNA (sRNA) _rprA_[134].