The second messenger signaling molecule cyclic di-AMP drives developmental cycle progression in Chlamydia trachomatis
- Department of Pathology, Microbiology, and Immunology, College of Medicine, University of Nebraska Medical Center, United States
Figures
Figure 1 with 1 supplement
Cyclic di-AMP accumulation is linked to secondary differentiation in C. trachomatis.
(A) Defining characteristics of chlamydial elementary bodies (EBs) and reticulate bodies (RBs). (B) A hypothetical model representing the correlation between c-di-AMP levels and the timing of secondary differentiation. The dashed line represents a threshold level of c-di-AMP needed to drive secondary differentiation in a given RB. hpi = hours post-infection. (C) Measurement of c-di-AMP concentrations in uninfected (-Ctr) and infected (+Ctr) HeLa cell lysates. For infected HeLa cells, C. trachomatis serovar L2 (434/Bu) transformed with an mCherry-encoding construct was infected into HeLa cells, and expression of mCherry was induced at 10 hpi with 5 nM anhydrotetracycline (aTc). All samples were harvested at 16 and 24 hpi. (D) A schematic diagram of the constructs used in this study. All constructs used are aTc-inducible as shown by the Ptet promoter. The location of the 6xH tag and the approximate location of the crRNA for the CRISPRi vectors are shown as well as the transmembrane domain of DacA. Diagram is not to scale. (E) Measurement of c-di-AMP concentrations in infected cell lysates from the strains shown in panel (D). C. trachomatis serovar L2 (434/Bu) transformed with the indicated constructs was infected into HeLa cells. At 10 hpi, expression of the construct was induced with 5 nM aTc, and the infected cells were harvested at 16 or 24 hpi. Levels of c-di-AMP in the supernatant were measured using ELISA. The left and right panels show the levels of c-di-AMP in the indicated strains at 16 and 24 hpi, respectively, on a log2 scale. For reference, 210=1024, 215=32,768, and 220=1,048,576. The dashed line in both graphs represents the c-di-AMP level of the 24 hpi mCherry-expressing control that is associated with EB production. N=3. *p<0.05, **p<0.001, NS: Not significant via two-sample equal variance t-test compared to the mCherry control.
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Figure 1—source data 1
Source data for c-di-AMP experiments shown in Figure 1.
- https://cdn.elifesciences.org/articles/104240/elife-104240-fig1-data1-v1.xlsx
Figure 1—figure supplement 1
The predicted transmembrane domains in DacA and YbbR.
Transmembrane domains in DacA (A) and YbbR (B) were predicted with TOPCONS (https://topcons.cbr.su.se/) (Tsirigos et al., 2015). The red and blue lines represent cytosolic and periplasmic domains, respectively.
Figure 2 with 6 supplements
Overexpression of DacA or DacA(D164N) is detrimental to the chlamydial developmental cycle.
HeLa cells were infected with C. trachomatis transformed with a plasmid encoding an anhydrotetracycline (aTc)-inducible DacA or DacA(D164N) (i.e., dacA or dacA(D164N), respectively; see Figure 1D). At 10 hpi, expression of the construct was induced or not with 5 nM aTc, and DNA and RNA samples were collected at 10, 14, and 24 hpi. Immunofluorescence analysis (IFA) and inclusion-forming units (IFU) samples were collected at 24 hpi. For IFA images, the green color represents chlamydial major outer membrane protein (MOMP), which shows the chlamydial cell morphology, and the red color represents DacA or DacA(D164N). (A &B) IFA images of the dacA(A) and dacA(D164N) (B) strains at 24 hpi. Shown are individual panels of a representative inclusion for the strains with DacA and MOMP labeling as well as the merged image. IFA images were acquired on a Zeiss AxioImager.Z2 equipped with an Apotome2 using a 100 X lens objective. Scale bar: 1 µm (A) or 2 µm (B). (C &D) Quantification of IFUs (C) and genomic DNA copy number (D) from uninduced and induced samples of dacA at 24 hpi. (E &F) Quantification of IFUs (E) and genomic DNA copy number (F) from uninduced and induced samples of dacA(D164N) at 24 hpi. (G &H) Quantification of transcripts by RT-qPCR for dacA, ybbR, euo, hctA, and omcB from uninduced and induced samples of dacA (G) and dacA(D164N) (H) UI = uninduced (i.e. -aTc); I=induced (i.e. +aTc) for all sample types. N=3. *p<0.05; **p<0.001 via two-sample equal variance t-test.
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Figure 2—source data 1
Source data for experiments shown in Figure 2.
- https://cdn.elifesciences.org/articles/104240/elife-104240-fig2-data1-v1.xlsx
Figure 2—figure supplement 1
The localization of DacA_6xH (A) or YbbR_6xH (B) in the first dividing cells.
C. trachomatis encoding DacA_6xH or YbbR_6xH was infected into HeLa cells. At 4 hpi, expression of the constructs was induced with 5 nM aTc. At 10.5 hpi, the infected cells were fixed with an aldehyde fixing solution (3.2% Formaldehyde, 0.022% Glutaraldehyde in 1 X PBS) for 2 min and permeabilized with 90% MeOH for 1 min. The images were acquired on a Zeiss Imager.Z2 equipped with an Apotome2 using a 100 X lens objective.
Figure 2—figure supplement 2
Inclusion area and cell diameter measurements of dacA and dacA(D164N) expressing strains.
(A) Inclusion area of the dacA strain. We measured the area of 56 inclusions (from n=3 replicates). (B) Cell diameter of the dacA strain. We measured the diameter of 142 (uninduced = UI) or 119 (induced = I) bacteria (from n=3 replicates). (C) Inclusion area of the dacA(D164N) strain. We measured the area of 54 inclusions (from n=3 replicates). (D) Cell diameter of the dacA(D164N) strain. We measured the diameter of 143 (UI) or 145 (I) bacteria (from n=3 replicates). The inclusion area and cell diameter were measured using Fiji software. **p<0.001 via two-sample equal variance t-test.
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Figure 2—figure supplement 2—source data 1
Source data for experiments shown in Figure 2—figure supplement 2.
- https://cdn.elifesciences.org/articles/104240/elife-104240-fig2-figsupp2-data1-v1.xlsx
Figure 2—figure supplement 3
Overexpression of YbbR_6xH does not affect the chlamydial developmental cycle.
HeLa cells were infected with C. trachomatis transformed with a plasmid encoding an anhydrotetracycline-inducible YbbR_6xH (i.e., ybbR_6xH; see Figure 1D). At 10 hpi, expression of the construct was induced or not with 5 nM aTc, and immunofluorescence assay (IFA), inclusion-forming unit (IFU), DNA, and RNA samples were collected. (A) IFA images of ybbR_6xH strain at 24 hpi. (B) Quantification of IFU from uninduced and induced samples at 24 hpi. (C) Quantification of genomic DNA copy number by qPCR in uninduced and induced samples. (D) Quantification of transcripts by RT-qPCR for dacA, ybbR, euo, hctA, and omcB. IFA images were acquired on a Zeiss AxioImager.Z2 equipped with an Apotome2 using a 100 X lens objective. Scale bar: 1 µm. UI = uninduced (i.e. -aTc); I=induced (i.e. +aTc) for all sample types. N=3. *p<0.05 via two-sample equal variance t-test.
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Figure 2—figure supplement 3—source data 1
Source data for experiments shown in Figure 2—figure supplement 3.
- https://cdn.elifesciences.org/articles/104240/elife-104240-fig2-figsupp3-data1-v1.xlsx
Figure 2—figure supplement 4
Overexpressed mCherry from the vector control does not affect elementary body (EB) progeny production during the chlamydial developmental cycle.
HeLa cells were infected with C. trachomatis transformed with a plasmid encoding an anhydrotetracycline-inducible mCherry. At 10 hpi, expression of the construct was induced or not with 5 nM aTc, and the EB samples (IFU - inclusion forming unit) were collected at 18, 20, 22, 24, 32, and 48 hpi.
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Figure 2—figure supplement 4—source data 1
Source data for experiments shown in Figure 2—figure supplement 4.
- https://cdn.elifesciences.org/articles/104240/elife-104240-fig2-figsupp4-data1-v1.xlsx
Figure 2—figure supplement 5
Overexpression of DacA_6xH is detrimental to the chlamydial developmental cycle.
HeLa cells were infected with C. trachomatis transformed with a plasmid encoding an anhydrotetracycline (aTc)-inducible DacA_6xH. At 10 hpi, expression of the construct was induced or not with 5 nM aTc, and DNA and RNA samples were collected at 10, 14, and 24 hpi. Immunofluorescence analysis (IFA) and inclusion-forming units (IFU) samples were collected at 24 hpi. (A) IFA images of the dacA_6xH strain at 24 hpi. Shown are individual panels for DacA_6xH and major outer membrane protein (MOMP) labeling as well as the merged image. IFA images were acquired on a Zeiss AxioImager.Z2 equipped with an Apotome2 using a 100 X lens objective. Scale bar: 1 µm. (B) Quantification of IFUs from uninduced and induced samples at 24 hpi. (C) Quantification of genomic DNA copy number by qPCR in uninduced and induced samples. (D) Quantification of transcripts by RT-qPCR for dacA, ybbR, euo, hctA, and omcB from uninduced and induced samples. UI = uninduced (i.e. -aTc); I=induced (i.e. +aTc) for all sample types. N=3. *p<0.05; **p<0.001 via two-sample equal variance t-test.
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Figure 2—figure supplement 5—source data 1
Source data for experiments shown in Figure 2—figure supplement 5.
- https://cdn.elifesciences.org/articles/104240/elife-104240-fig2-figsupp5-data1-v1.xlsx
Figure 2—figure supplement 6
Quantification of transcripts by RT-qPCR for dacA, ybbR, euo, hctA, and omcB from dacA-KD and dacA_6xH strains cultured in STING-KO HeLa cells.
STING-KO HeLa cells were infected with C. trachomatis transformed with a plasmid encoding an anhydrotetracycline (aTc)-inducible (A) DacA_6xH and/or (B). CRISPRi-dCas12 system targeting the dacA promoter. At 10 hpi, expression of the construct was induced or not with 5 nM aTc, and DNA and RNA samples were collected at 10, 14, and 24 hpi.
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Figure 2—figure supplement 6—source data 1
Source data for experiments shown in Figure 2—figure supplement 6.
- https://cdn.elifesciences.org/articles/104240/elife-104240-fig2-figsupp6-data1-v1.xls
Figure 3
Overexpression of ΔTMDacA or ΔTMDacA(D164N) does not alter the chlamydial developmental cycle.
HeLa cells were infected with C. trachomatis transformed with a plasmid encoding an anhydrotetracycline (aTc)-inducible ΔTMDacA or ΔTMDacA(D164N) (i.e. ΔTMdacA and ΔTMdacA (D164N); see Figure 1D). At 10 hpi, expression of the construct was induced or not with 5 nM aTc, and DNA and RNA samples were collected at 10, 14, and 24 hpi. Immunofluorescence analysis (IFA) and inclusion-forming units (IFU) samples were collected at 24 hpi. For IFA images, the green color represents chlamydial major outer membrane protein (MOMP), which shows the chlamydial cell morphology, and the red color represents ΔTMDacA or ΔTMDacA(D164N). (A & B) IFA images of ΔTMdacA(A) and ΔTMdacA(D164N)(B) strains at 24 hpi. Shown are individual panels of a representative inclusion for the strains with DacA and MOMP labeling as well as the merged image. IFA images were acquired on a Zeiss AxioImager.Z2 equipped with an Apotome2 using a 100 X lens objective. Scale bar: 2 µm. (C & D) Quantification of IFUs (C) and genomic DNA copy number (D) from uninduced and induced samples of ΔTMdacA at 24 hpi. (E & F) Quantification of IFUs (E) and genomic DNA copy number (F) from uninduced and induced samples of ΔTMdacA(D164N). (G & H) Quantification of transcripts by RT-qPCR for dacA, ybbR, euo, hctA, and omcB from uninduced and induced samples of ΔTMdacA (G) and ΔTMdacA(D164N) (H) UI = uninduced (i.e. -aTc); I=induced (i.e. +aTc) for all sample types. N=3. *p<0.05 via two-sample equal variance t-test.
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Figure 3—source data 1
Source data for experiments shown in Figure 3.
- https://cdn.elifesciences.org/articles/104240/elife-104240-fig3-data1-v1.xlsx
Figure 4 with 1 supplement
CRISPRi-mediated dacA-ybbR knockdown displays reduced levels of transcripts for late genes.
HeLa cells were infected with C. trachomatis transformed with a plasmid encoding an anhydrotetracycline (aTc)-inducible CRISPRi-dCas12 system targeting the dacA promoter (dacA-KD) or dacA-KD system and DacA/YbbR_6xH (i.e., dacA-KDcom; see Figure 1D). At 10 hpi, knockdown was induced or not with 5 nM aTc, and DNA and RNA samples were collected at 10, 14, and 24 hpi. Immunofluorescence analysis (IFA) and inclusion-forming units (IFU) samples were collected at 24 hpi. (A & B) IFA images of the dacA-KD (A) and dacA-KDcom (B) strains at 24 hpi. Shown are individual panels of a representative inclusion for the strains for dCas12, YbbR_6xH, and major outer membrane protein (MOMP) labeling as well as the merged image. IFA images were acquired on a Zeiss AxioImager.Z2 equipped with an Apotome2 using a 100 X lens objective. Scale bar: 2(A) or 1(B) µm. (C & D) Quantification of IFUs (C) and genomic DNA copy number (D) from uninduced and induced samples of dacA-KD at 24 hpi. (E and F) Quantification of IFUs (E) and genomic DNA copy number (F) in uninduced and induced samples of dacA-KDcom. (G & H) Quantification of transcripts by RT-qPCR for dacA, ybbR, euo, hctA, and omcB from uninduced and induced samples of dacA-KD (G) and dacA-KDcom (H) UI = uninduced (i.e. -aTc); I=induced (i.e. +aTc) for all sample types. N=3. *p<0.05; **p<0.001 via two-sample equal variance t-test.
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Figure 4—source data 1
Source data for experiments shown in Figure 4.
- https://cdn.elifesciences.org/articles/104240/elife-104240-fig4-data1-v1.xlsx
Figure 4—figure supplement 1
Inclusion area and cell diameter measurements of dacA-KD and dacA-KDcom.
(A) Inclusion area of the dacA-KD strain. We measured the area of 30 (uninduced = UI) and 29 (induced = I) inclusions (from n=3 replicates). (B) Cell diameter of the dacA-KD strain. We measured the diameter of 126 (UI) or 129 (induced = I) bacteria (from n=3 replicates). (C) Inclusion area of the dacA-KDcom strain. We measured the area of 62 (UI) and 60 (I) inclusions (from n=3 replicates). (D) Cell diameter of the dacA-KDcom strain. We measured the diameter of 149 (UI) and 142 (I) bacteria (from n=3 replicates). The inclusion area and cell diameter were measured using Fiji software. **p<0.001 via two-sample equal variance t-test.
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Figure 4—figure supplement 1—source data 1
Source data for experiments shown in Figure 4—figure supplement 1.
- https://cdn.elifesciences.org/articles/104240/elife-104240-fig4-figsupp1-data1-v1.xlsx
Figure 5 with 2 supplements
Overexpression of DacA and YbbR_6xH prematurely increases hctA transcript levels.
HeLa cells were infected with C. trachomatis transformed with an aTc-inducible plasmid encoding wild-type DacA/YbbR_6xH or DacA(D164N)/YbbR_6xH (i.e., dacAop and dacAopMut, respectively; see Figure 1D). At 10 hpi, expression of the constructs was induced or not with 5 nM aTc, and DNA and RNA samples were collected at 10, 14, and 24 hpi. Immunofluorescence analysis (IFA) samples were collected at 24 hpi. (A &B). IFA images of the dacAop (A) and dacAopMut (B) at 24 hpi. Shown are individual panels of a representative inclusion for the strains for DacA and YbbR_6xH as well as the merged image with major outer membrane protein (MOMP) labeling. The arrowheads represent the co-localization of DacA and YbbR. IFA images were acquired on a Zeiss AxioImager.Z2 equipped with an Apotome2 using a 100 X lens objective. Scale bar: 2 µm. (C & D) Quantification of IFUs (C) and genomic DNA copy number (D) from uninduced and induced samples of dacAop at 24 hpi. (E & F). Quantification of IFUs (E) and genomic DNA copy number (F) from uninduced and induced samples of dacAopMut. (G & H). Quantification of transcripts by RT-qPCR for dacA, ybbR, euo, hctA, and omcB from uninduced and induced samples of dacAop (G) and dacAopMut (H). UI = uninduced (i.e. -aTc); I=induced (i.e. +aTc) for all sample types. N=3. *p<0.05 two-sample equal variance t-test.
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Figure 5—source data 1
Source data for experiments shown in Figure 5.
- https://cdn.elifesciences.org/articles/104240/elife-104240-fig5-data1-v1.xlsx
Figure 5—figure supplement 1
Inclusion area and cell diameter measurements of dacAop and dacAopMut.
(A) Inclusion area of the dacAop strain. We measured the area of 52 (uninduced = UI) and 58 (induced = I) inclusions (from n=3 replicates). (B) Cell diameter of the dacAop strain. We measured the diameter of 125 (UI) or 131 (induced = I) bacteria (from n=3 replicates). (C) Inclusion area of the dacAopMut strain. We measured the area of 50 (UI) and 52 (I) inclusions (from n=3 replicates). (D). Cell diameter of the dacAopMut strain. We measured the diameter of 124 (UI) and 150 (I) bacteria (from n=3 replicates). The inclusion area and cell diameter were measured using Fiji software. **p<0.001 via two-sample equal variance t-test.
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Figure 5—figure supplement 1—source data 1
Source data for experiments shown in Figure 5—figure supplement 1.
- https://cdn.elifesciences.org/articles/104240/elife-104240-fig5-figsupp1-data1-v1.xlsx
Figure 5—figure supplement 2
Phenotypic characterization of a dacAop overexpression construct encoding spectinomycin resistance (SpcR).
HeLa cells were infected with C. trachomatis transformed with a dacAop overexpression construct encoding SpcR. At 10 hpi, expression of the construct was induced or not with 5 nM aTc, and DNA and RNA samples were collected at 10, 14, and 24 hpi. Immunofluorescence analysis (IFA) samples were collected at 24 hpi. (A) IFA images of the dacAop strain at 24 hpi. Shown are individual panels for DacA, YbbR_6xH, and major outer membrane protein (MOMP) labeling as well as the merged image. (B) Quantification of genomic DNA copy number by qPCR in uninduced and induced samples. (C) Quantification of transcripts by RT-qPCR for dacA, ybbR, euo, hctA, and omcB. IFA images were acquired on a Zeiss AxioImager.Z2 equipped with an Apotome2 using a 100 X lens objective. Scale bar: 2 µm. UI = uninduced (i.e. -aTc); I=induced (i.e. +aTc) for all sample types. N=3. *p<0.05 via two-sample equal variance t-test.
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Figure 5—figure supplement 2—source data 1
Source data for experiments shown in Figure 5—figure supplement 2.
- https://cdn.elifesciences.org/articles/104240/elife-104240-fig5-figsupp2-data1-v1.xlsx
Figure 6
The levels of c-di-AMP are correlated with late gene transcripts.
RNA sequencing was performed from HeLa cells infected with either the dacAop or dacA-KD strains. RNA samples were collected at 16 hpi for dacAop and 24 hpi for dacA-KD after inducing expression of the relevant constructs at 10 hpi. Shown is a volcano plot of the RNA sequencing results with the vertical dashed lines indicating a twofold change in transcript levels as compared to the respective uninduced control for the given strain and the horizontal lines indicating a p-value of 0.05. The plot was made using GraphPad Prism software. Green spots represent genes demonstrating a statistically significant twofold change in transcription levels between uninduced and induced samples. Red dots represent genes with significant changes in transcript levels less than twofold. Blue dots represent genes not significantly different but more than twofold changed between the conditions. Black dots represent genes not significantly different and less than twofold changed between the conditions. See also Table 1 and Supplementary file 1 and Supplementary file 2 for more details.
Figure 7
Elementary body (EB) production is induced by high levels of c-di-AMP.
(A–C) HeLa cells were infected with the chlamydial transformants dacAop (A), dacA-KD (B), or dacAopMut (C). Expression of the constructs was induced or not with 5 nM aTc at 10 hpi. At 18, 20, 22, 24, 32, and 48 hpi, infected cell lysates were harvested for inclusion-forming unit (IFU) quantification. UI = uninduced (i.e. -aTc); I=induced (i.e. +aTc) for all sample types. #: Non-detected from the induced samples. N=3. *p<0.05; **p<0.001 via two-sample equal variance t-test.
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Figure 7—source data 1
Source data for experiments shown in Figure 7.
- https://cdn.elifesciences.org/articles/104240/elife-104240-fig7-data1-v1.xlsx
Figure 8
A working model illustrating how c-di-AMP impacts the onset of secondary differentiation in C. trachomatis.
(A) During reticulate body (RB) division, there is a gradient of c-di-AMP that forms between the mother and daughter cell. The lighter blue represents lower levels of c-di-AMP, the darker blue represents higher levels of c-di-AMP. In the depicted scenario, the daughter cell is able to divide again, having not reached a critical threshold of c-di-AMP. In contrast, the mother cell accumulates sufficient c-di-AMP to trigger secondary differentiation to an EB. (B) A model illustrating how c-di-AMP levels impact secondary differentiation progression. Shown is the ‘normal’ condition of wild-type bacteria, bacteria overexpressing DacA and YbbR (higher cyclic di-AMP), and bacteria expressing the CRISPRi system targeting the dacA promoter (lower cyclic di-AMP). Depending on the amount of c-di-AMP produced, secondary differentiation can either be triggered earlier or later as shown. The black dots represent EBs; the bigger and blue-colored circles show the RB cells. The brightness of blue reflects the c-di-AMP level.
Tables
Table 1
Genes impacted by cyclic di-AMP levels.
| Canonical late genes | Fold change | |||||
|---|---|---|---|---|---|---|
| Gene ID | Ctr D ORF | Name | Protein names | dacAop_OE | dacA-KD | Reference |
| CTL0112 | CT743 | hctA | Histone H1-like protein HC1 | 6.70 | –3.47 | Belland et al., 2003; Fahr et al., 1995 |
| CTL0302 | CT046 | hct2 | Histone H1-like protein HC2 | 3.52 | –5.96 | Belland et al., 2003; Yu et al., 2006 |
| CTL0336 | CT080 | ltuB | Late transcription unit B protein | 4.00 | –3.61 | Belland et al., 2003; Fahr et al., 1995 |
| CTL0700 | CT441 | tsp | Carboxy-terminal processing protease | 6.18 | –6.62 | Yu et al., 2006 |
| CTL0702 | CT443 | omcB | Large cysteine-rich periplasmic protein | 5.00 | –4.04 | Belland et al., 2003; Soules et al., 2020b |
| CTL0703 | CT444 | omcA | Small cysteine-rich outer membrane protein | 5.93 | –3.71 | Belland et al., 2003; Soules et al., 2020b |
| CTL0716 | CT456 | tarp | Translocated actin-recruiting phosphoprotein | 6.06 | –5.97 | Hatch and Ouellette, 2023; Soules et al., 2020b |
| Membrane organization associated | Fold change | |||||
| Gene ID | Ctr D ORF | Name | Protein Names | dacAop_OE | dacA-KD | Reference |
| CTL0082 | CT713 | ompB | Outer membrane protein B | 2.73 | –2.15 | Nicholson et al., 2003 |
| CTL0248 | CT869 | pmpE | Polymorphic outer membrane protein | 1.99 | –2.04 | Hatch and Ouellette, 2023; Nicholson et al., 2003 |
| CTL0249 | CT870 | pmpF | Polymorphic outer membrane protein | 3.03 | –2.23 | Hatch and Ouellette, 2023; Nicholson et al., 2003 |
| CTL0250 | CT871 | pmpG | Polymorphic outer membrane protein | 2.03 | –1.55 | Belland et al., 2003; Nicholson et al., 2003 |
| CTL0429 | CT177 | dsbA | Disulfide bond chaperone | 2.79 | –2.47 | Hatch and Ouellette, 2023 |
| CTL0610 | CT356 | dsbH | Thioredox_DsbH domain-containing protein | 3.07 | –2.11 | Belland et al., 2003; Nicholson et al., 2003 |
| CTL0670 | CT413 | pmpB | Polymorphic outer membrane protein | 4.17 | –1.74 | Hatch and Ouellette, 2023; Nicholson et al., 2003 |
| Gene Regulation associated | Fold change | |||||
| Gene ID | Ctr D ORF | Name | Protein Names | dacAop_OE | dacA-KD | Reference |
| CTL0044 | CT675 | mcsB | Protein-arginine kinase | 3.52 | –2.21 | Hatch and Ouellette, 2023 |
| CTL0045 | CT676 | mcsA | UVR domain-containing protein | 2.53 | –2.92 | Hatch and Ouellette, 2023 |
| CTL0727 | CT467 | atoS | Two component regulator, histidine kinase | 3.33 | –1.80 | Hatch and Ouellette, 2023 |
| CTL0728 | CT468 | atoC | Two-component system response regulator | 7.95 | –2.78 | |
| CTL0894 | CT630 | chxR | Atypical response regulator protein ChxR | 3.68 | –2.93 | Yang et al., 2017 |
| Glycogen synthesis | Fold change | |||||
| Gene ID | Ctr D ORF | Name | Protein Names | dacAop_OE | dacA-KD | Reference |
| CTL0167 | CT798 | glgA | Glycogen synthase | 9.58 | –2.86 | Belland et al., 2003; Nicholson et al., 2003 |
| CTL0342 | CT087 | malQ | 4-alpha-glucanotransferase | 9.43 | –4.29 | Hefty and Stephens, 2007 |
| CTL0500 | CT248 | glgP | Alpha-1,4 glucan phosphorylase | 2.70 | –1.71 | Nicholson et al., 2003 |
| Type III Secretion System | Fold change | |||||
| Gene ID | Ctr D ORF | Name | Protein Names | dacAop_OE | dacA-KD | Reference |
| CTL0041 | CT672 | sctQ | Type III secretion component, basal body | 1.93 | –1.76 | Hefty and Stephens, 2007 |
| CTL0043 | CT674 | cdsC | Type III secretion structural protein | 1.79 | –1.56 | Hefty and Stephens, 2007 |
| CTL0343 | CT088 | scc1 | Type III secretion chaperone | 4.48 | –3.78 | Hefty and Stephens, 2007 |
| CTL0824 | CT561 | sctL | Type III secretion system protein | 2.18 | –1.75 | Hatch and Ouellette, 2023 |
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OE = overexpression.
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KD = knockdown.
Additional files
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MDAR checklist
- https://cdn.elifesciences.org/articles/104240/elife-104240-mdarchecklist1-v1.docx
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Supplementary file 1
Generalized representation of sequencing efficiency.
- https://cdn.elifesciences.org/articles/104240/elife-104240-supp1-v1.xlsx
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Supplementary file 2
RNA sequencing results.
- https://cdn.elifesciences.org/articles/104240/elife-104240-supp2-v1.xlsx
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Supplementary file 3
List of plasmids and primers used in the study.
- https://cdn.elifesciences.org/articles/104240/elife-104240-supp3-v1.docx
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The second messenger signaling molecule cyclic di-AMP drives developmental cycle progression in Chlamydia trachomatis
eLife 14:RP104240.
https://doi.org/10.7554/eLife.104240.4
