The reliability of colorimetry is precise(ly) as expected.

Figure S6. Gene ontology (GO) analysis of the differentially expressed genes between C1 and C2. (JPG 403 kb

The effect of putative anti-σ factor Tt-TolB on the activity of Tt-RpoE1.

<p>(<b>A</b>). The effect of Tt-TolB on the interaction between Tt-RpoE1 and fork-junction structure promoter DNA (T+1/B−10). The indicated proteins were added (+) in an EMSA reaction at a concentration of 5 µM. The solid arrow indicates the supershifted complex formed by Tt-RpoE1, Tt-TolB, and the promoter, and the open arrow indicates the complex formed by Tt-RpoE1 and the promoter. (<b>B</b>). The effect of Tt-TolB on <i>in vitro</i> transcription of Tt-RpoE1. Lane 1. Tt-TolB was added into the transcription system at the same time with Tt-RpoE1. Lane 2. Tt-TolB was added into the transcription system after Tt-RpoE1, <i>E.coli</i> core RNAP and promoter DNA being incubated (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0040885#s4" target="_blank">materials and methods</a>). Lane C, the <i>in vitro</i> transcription system without Tt-TolB. The solid arrow indicates the products of transcription.</p

Partial oligonucleotides used in this study.

*<p>Restriction sites are underlined.</p

Protein Dynamic Landscape during Mouse Mammary Gland Development from Virgin to Pregnant, Lactating, and Involuting Stages

The mammary gland undergoes significant physiological changes as it undergoes a transition from virgin to pregnancy, lactation, and involution. However, the dynamic role of proteins in regulating these processes during mouse mammary gland development has not been thoroughly explored. In this study, we collected mouse mammary gland tissues from mature virgins aged 8–10 weeks (V), day 16 of pregnancy (P16d), day 12 of lactation (L12d), day 1 of forced weaning (FW 1d), and day 3 of forced weaning (FW 3d) stages for analysis using DIA-based quantitative proteomics technology. A total of 3,312 proteins were identified, of which 843 were DAPs that were categorized into nine clusters based on their abundance changes across developmental stages. Notably, DAPs in cluster 2, which peaked at the L12d stage, were primarily associated with mammary gland development and lactation. The protein–protein interaction network revealed that the epidermal growth factor (EGF) was central to this cluster. Our study provides a comprehensive overview of the mouse mammary gland development proteome and identifies some important proteins, such as EGF, Janus kinase 1 (JAK1), and signal transducer and activator of transcription 6 (STAT6) that may serve as potential targets for future research to provide guidelines for a deeper understanding of the developmental biology of mammary glands

Protein Dynamic Landscape during Mouse Mammary Gland Development from Virgin to Pregnant, Lactating, and Involuting Stages

The mammary gland undergoes significant physiological changes as it undergoes a transition from virgin to pregnancy, lactation, and involution. However, the dynamic role of proteins in regulating these processes during mouse mammary gland development has not been thoroughly explored. In this study, we collected mouse mammary gland tissues from mature virgins aged 8–10 weeks (V), day 16 of pregnancy (P16d), day 12 of lactation (L12d), day 1 of forced weaning (FW 1d), and day 3 of forced weaning (FW 3d) stages for analysis using DIA-based quantitative proteomics technology. A total of 3,312 proteins were identified, of which 843 were DAPs that were categorized into nine clusters based on their abundance changes across developmental stages. Notably, DAPs in cluster 2, which peaked at the L12d stage, were primarily associated with mammary gland development and lactation. The protein–protein interaction network revealed that the epidermal growth factor (EGF) was central to this cluster. Our study provides a comprehensive overview of the mouse mammary gland development proteome and identifies some important proteins, such as EGF, Janus kinase 1 (JAK1), and signal transducer and activator of transcription 6 (STAT6) that may serve as potential targets for future research to provide guidelines for a deeper understanding of the developmental biology of mammary glands

Protein Dynamic Landscape during Mouse Mammary Gland Development from Virgin to Pregnant, Lactating, and Involuting Stages

The mammary gland undergoes significant physiological changes as it undergoes a transition from virgin to pregnancy, lactation, and involution. However, the dynamic role of proteins in regulating these processes during mouse mammary gland development has not been thoroughly explored. In this study, we collected mouse mammary gland tissues from mature virgins aged 8–10 weeks (V), day 16 of pregnancy (P16d), day 12 of lactation (L12d), day 1 of forced weaning (FW 1d), and day 3 of forced weaning (FW 3d) stages for analysis using DIA-based quantitative proteomics technology. A total of 3,312 proteins were identified, of which 843 were DAPs that were categorized into nine clusters based on their abundance changes across developmental stages. Notably, DAPs in cluster 2, which peaked at the L12d stage, were primarily associated with mammary gland development and lactation. The protein–protein interaction network revealed that the epidermal growth factor (EGF) was central to this cluster. Our study provides a comprehensive overview of the mouse mammary gland development proteome and identifies some important proteins, such as EGF, Janus kinase 1 (JAK1), and signal transducer and activator of transcription 6 (STAT6) that may serve as potential targets for future research to provide guidelines for a deeper understanding of the developmental biology of mammary glands

Protein Dynamic Landscape during Mouse Mammary Gland Development from Virgin to Pregnant, Lactating, and Involuting Stages

The mammary gland undergoes significant physiological changes as it undergoes a transition from virgin to pregnancy, lactation, and involution. However, the dynamic role of proteins in regulating these processes during mouse mammary gland development has not been thoroughly explored. In this study, we collected mouse mammary gland tissues from mature virgins aged 8–10 weeks (V), day 16 of pregnancy (P16d), day 12 of lactation (L12d), day 1 of forced weaning (FW 1d), and day 3 of forced weaning (FW 3d) stages for analysis using DIA-based quantitative proteomics technology. A total of 3,312 proteins were identified, of which 843 were DAPs that were categorized into nine clusters based on their abundance changes across developmental stages. Notably, DAPs in cluster 2, which peaked at the L12d stage, were primarily associated with mammary gland development and lactation. The protein–protein interaction network revealed that the epidermal growth factor (EGF) was central to this cluster. Our study provides a comprehensive overview of the mouse mammary gland development proteome and identifies some important proteins, such as EGF, Janus kinase 1 (JAK1), and signal transducer and activator of transcription 6 (STAT6) that may serve as potential targets for future research to provide guidelines for a deeper understanding of the developmental biology of mammary glands

Identification and Characterization of the Cognate Anti-Sigma Factor and Specific Promoter Elements of a <em>T. tengcongensis</em> ECF Sigma Factor

<div><p>Extracytoplasmic function (ECF) σ factors, the largest group of alternative σ factors, play important roles in response to environmental stresses. Tt-RpoE1 is annotated as an ECF σ factor in <em>Thermoanaerobacter tengcongensis</em>. In this study, we revealed that the <em>Tt-tolB</em> gene located downstream of the <em>Tt-rpoE1</em> gene encoded the cognate anti-σ factor, which could inhibit the transcription activity of Tt-RpoE1 by direct interaction with Tt-RpoE1 via its N-terminal domain. By <em>in vitro</em> transcription assay, the auto-regulation ability of Tt-RpoE1 was determined, and band shift assay showed that Tt-RpoE1 preferred to bind a fork-junction promoter DNA. With truncation or base-specific scanning mutations, the contribution of the nucleotides in −35 and −10 regions to interaction between Tt-RpoE1 and promoter DNA was explored. The promoter recognition pattern of Tt-RpoE1 was determined as 5′ tGTTACN₁₆CGTC 3′, which was further confirmed by <em>in vitro</em> transcription assays. This result showed that the Tt-RpoE1-recognized promoter possessed a distinct −10 motif (−13CGTC−10) as the recognition determinant, which is distinguished from the −10 element recognized by σ⁷⁰. Site-directed mutagenesis in Region 2.4 of Tt-RpoE1 indicated that the “D” residue of DXXR motif was responsible for recognizing the −12G nucleotide. Our results suggested that distinct −10 motif may be an efficient and general strategy used by ECF σ factors in adaptive response regulation of the related genes.</p> </div

The effect of scanning substitutions in the −35 and −10 regions on the activity of Tt-RpoE1.

<p>(<b>A & B</b>). EMSA results of substitutions in the −35 (<b>A</b>) and −10 (<b>B</b>) regions. The D−5/−10 or D−5/−12 fork-junction structure of promoter DNA was used as the parental probe respectively. The substitutions were made both on the top and opposite positions of the bottom strands in the duplex part as indicated. (<b>C</b>). The effect of a subset of substitutions in the −10 region on <i>in vitro</i> transcription of Tt-RpoE1.</p

Protein Dynamic Landscape during Mouse Mammary Gland Development from Virgin to Pregnant, Lactating, and Involuting Stages

The mammary gland undergoes significant physiological changes as it undergoes a transition from virgin to pregnancy, lactation, and involution. However, the dynamic role of proteins in regulating these processes during mouse mammary gland development has not been thoroughly explored. In this study, we collected mouse mammary gland tissues from mature virgins aged 8–10 weeks (V), day 16 of pregnancy (P16d), day 12 of lactation (L12d), day 1 of forced weaning (FW 1d), and day 3 of forced weaning (FW 3d) stages for analysis using DIA-based quantitative proteomics technology. A total of 3,312 proteins were identified, of which 843 were DAPs that were categorized into nine clusters based on their abundance changes across developmental stages. Notably, DAPs in cluster 2, which peaked at the L12d stage, were primarily associated with mammary gland development and lactation. The protein–protein interaction network revealed that the epidermal growth factor (EGF) was central to this cluster. Our study provides a comprehensive overview of the mouse mammary gland development proteome and identifies some important proteins, such as EGF, Janus kinase 1 (JAK1), and signal transducer and activator of transcription 6 (STAT6) that may serve as potential targets for future research to provide guidelines for a deeper understanding of the developmental biology of mammary glands