| Immunostaining of surface-bound bacteria | On the following day, HeLa cells were then infected with invasin-expressing E. coli at an MOI of 200 by plate rotation at an rpm of 50, in a 37°C, 5% C02 humidified incubator for 90min. |
| Immunostaining of surface-bound bacteria | After infection, HeLa cell were washed twice with phosphoate buffered saline (PBS) and fixed with 3% formaldehyde/ 0.025% glutaraldehyde at room temperature for 10 min. |
| Immunostaining of surface-bound bacteria | HeLa cells were then blocked with 5% BSA in PBS for 30 min at room temperature and stained with anti-LPS E. coli (Abcam). |
| Supporting Information | HeLa cells were co-incubated with E. coli |
| Supporting Information | HeLa cells incubated With either control antibody (actin) or antibody against Bl-integrin along With fluorescent secondary antibody (Alexa647). |
| Supporting Information | HeLa cells , infected with E. coli expressing invasin and GFP at an MOI of 200 for 90 minutes, were washed with PBS twice, fixed with paraformaldehyde, and immunostained with an anti-LPS antibody to identify bacterial cell wall (red color) and Hoechst to identify DNA (blue color). |
| The probability of invasin-mediated uptake is invariant | After co-culture of increasing amounts of GFP-expressing E. coli harboring a plasmid permitting arabinose-inducible control of invasin (pBACr-Aralnv) with HeLa cells , flow cytometry was conducted to obtain data shown in Fig 3A. |
| The probability of invasin-mediated uptake is invariant | In the absence of induction, bacteria carrying pBACr-AraIm/ cannot infect host cells, which was reported previously [27] and observed by us by co-incubating HeLa cells with uninduced bacteria (S6A—S6C Fig). |
| Variability in invasin-mediated bacterial uptake | In each experiment, the engineered bacteria were co-cultured with HeLa cells for 90 minutes in well-mixed conditions to mitigate the effects of heterogeneity in the bacterial population. |
| Variability in invasin-mediated bacterial uptake | Consistent with observations reported in the literature [18,22], fluorescence microscopy confirmed drastic cell-cell variability in bacterial uptake by HeLa cells (Fig 1A). |
| Variability in invasin-mediated bacterial uptake | However, immunolabeling experiments revealed a relatively narrow, unimodal distribution of Bl-integrins across HeLa cells (SID Fig). |
| Introduction | The attenuation of the initial response of delayed IEGs to EGF was also shown to depend on negative feedback through de novo transcription in HeLa cells [10], but it is unknown whether negative feedback plays roles in other cell types or under other stimuli. |
| Kinetics and chromatin features underlying IEG induction | This set of delayed early genes has been shown to be activated in waves following FOS, JUN and EGRl expression in HeLa cells [10]. |
| Kinetics and chromatin features underlying IEG induction | This is consistent with reports that NAB2 represses EGR1 and thereby attenuates the immediate-early response in HeLa cells stimulated with EGF [10]. |