Limits...
The Multiple DSF-family QS Signals are Synthesized from Carbohydrate and Branched-chain Amino Acids via the FAS Elongation Cycle.

Zhou L, Yu Y, Chen X, Diab AA, Ruan L, He J, Wang H, He YW - Sci Rep (2015)

Bottom Line: Furthermore, our biochemical analyses show that the key DSF synthase RpfF has both thioesterase and dehydratase activities, and uses 3-hydroxydedecanoyl-ACP as a substrate to produce BDSF.Finally, our results show that the classic fatty acid synthesis elongation cycle is required for the biosynthesis of DSF-family signals.Taken all together, these findings establish a general biosynthetic pathway for the DSF-family quorum sensing signals.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Microbial Metabolism, School of Life Sciences &Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China.

ABSTRACT
Members of the diffusible signal factor (DSF) family are a novel class of quorum sensing (QS) signals in diverse Gram-negative bacteria. Although previous studies have identified RpfF as a key enzyme for the biosynthesis of DSF family signals, many questions in their biosynthesis remain to be addressed. In this study with the phytopathogen Xanthomonas campestris pv. campestris (Xcc), we show that Xcc produces four DSF-family signals (DSF, BDSF, CDSF and IDSF) during cell culture, and that IDSF is a new functional signal characterized as cis-10-methyl-2-dodecenoic acid. Using a range of defined media, we further demonstrate that Xcc mainly produces BDSF in the presence of carbohydrates; leucine and valine are the primary precursor for DSF biosynthesis; isoleucine is the primary precursor for IDSF biosynthesis. Furthermore, our biochemical analyses show that the key DSF synthase RpfF has both thioesterase and dehydratase activities, and uses 3-hydroxydedecanoyl-ACP as a substrate to produce BDSF. Finally, our results show that the classic fatty acid synthesis elongation cycle is required for the biosynthesis of DSF-family signals. Taken all together, these findings establish a general biosynthetic pathway for the DSF-family quorum sensing signals.

No MeSH data available.


Related in: MedlinePlus

The effects of leucine and valine on DSF biosynthesis.(a) Effect of total amino acids and total non-branched-chain amino acids on DSF-family signals production. XYSA: XYS medium supplemented with 100 μM total amino acids; XYSN: XSY medium supplemented with 100 μM non-branched-chain amino acids. (b) HPLC analysis of DSF and BDSF production by the ΔrpfC strain in media XYS (as indicated in black line), XYSL (red line) and XYSV (blue line) at 24 h after inoculation. XYSL: XYS medium supplemented with 50 μM leucine. XYSV: XYS medium supplemented with 350 μM valine. (c) The dose–response curve between leucine or valine concentration and DSF production. (d) DSF production time course of the ΔrpfC strain in media XYS, XYSL, and XYSV. Data are means ± one standard deviation of three independent assays. Different letters indicate significant differences between treatments (LSD at P = 0.05).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4542539&req=5

f3: The effects of leucine and valine on DSF biosynthesis.(a) Effect of total amino acids and total non-branched-chain amino acids on DSF-family signals production. XYSA: XYS medium supplemented with 100 μM total amino acids; XYSN: XSY medium supplemented with 100 μM non-branched-chain amino acids. (b) HPLC analysis of DSF and BDSF production by the ΔrpfC strain in media XYS (as indicated in black line), XYSL (red line) and XYSV (blue line) at 24 h after inoculation. XYSL: XYS medium supplemented with 50 μM leucine. XYSV: XYS medium supplemented with 350 μM valine. (c) The dose–response curve between leucine or valine concentration and DSF production. (d) DSF production time course of the ΔrpfC strain in media XYS, XYSL, and XYSV. Data are means ± one standard deviation of three independent assays. Different letters indicate significant differences between treatments (LSD at P = 0.05).

Mentions: To investigate whether amino acids influence the production of DSF-family signals, total amino acids or total amino acids without branched-chain amino acids (BCAAs) at 100 μM were added separately to medium XYS. The resulting media, XYSA and XYSN respectively, supported the growth of the ΔrpfC strain to a maximum OD600 of 1.2. In XYSA medium, the ΔrpfC strain produced three DSF-family signals, DSF (0.9 μM or 2.0 × 10−3 μmol/109 cells), BDSF (2.8 μM or 6.2 × 10−3 μmol/109 cells) and IDSF (0.08 μM or 0.18 × 10−3 μmol/109 cells) (Fig. 3a). However, the ΔrpfC strain grown in XSYN medium produced only two DSF-family signals, DSF at a reduced level (0.22 μM or 0.48 × 10−3 μmol/109 cells), and BDSF (2.9 μM or 6.4 × 10−3 μmol/109 cells) (Fig. 3a). These results suggest that BCAAs are important for both DSF and IDSF biosynthesis.


The Multiple DSF-family QS Signals are Synthesized from Carbohydrate and Branched-chain Amino Acids via the FAS Elongation Cycle.

Zhou L, Yu Y, Chen X, Diab AA, Ruan L, He J, Wang H, He YW - Sci Rep (2015)

The effects of leucine and valine on DSF biosynthesis.(a) Effect of total amino acids and total non-branched-chain amino acids on DSF-family signals production. XYSA: XYS medium supplemented with 100 μM total amino acids; XYSN: XSY medium supplemented with 100 μM non-branched-chain amino acids. (b) HPLC analysis of DSF and BDSF production by the ΔrpfC strain in media XYS (as indicated in black line), XYSL (red line) and XYSV (blue line) at 24 h after inoculation. XYSL: XYS medium supplemented with 50 μM leucine. XYSV: XYS medium supplemented with 350 μM valine. (c) The dose–response curve between leucine or valine concentration and DSF production. (d) DSF production time course of the ΔrpfC strain in media XYS, XYSL, and XYSV. Data are means ± one standard deviation of three independent assays. Different letters indicate significant differences between treatments (LSD at P = 0.05).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4542539&req=5

f3: The effects of leucine and valine on DSF biosynthesis.(a) Effect of total amino acids and total non-branched-chain amino acids on DSF-family signals production. XYSA: XYS medium supplemented with 100 μM total amino acids; XYSN: XSY medium supplemented with 100 μM non-branched-chain amino acids. (b) HPLC analysis of DSF and BDSF production by the ΔrpfC strain in media XYS (as indicated in black line), XYSL (red line) and XYSV (blue line) at 24 h after inoculation. XYSL: XYS medium supplemented with 50 μM leucine. XYSV: XYS medium supplemented with 350 μM valine. (c) The dose–response curve between leucine or valine concentration and DSF production. (d) DSF production time course of the ΔrpfC strain in media XYS, XYSL, and XYSV. Data are means ± one standard deviation of three independent assays. Different letters indicate significant differences between treatments (LSD at P = 0.05).
Mentions: To investigate whether amino acids influence the production of DSF-family signals, total amino acids or total amino acids without branched-chain amino acids (BCAAs) at 100 μM were added separately to medium XYS. The resulting media, XYSA and XYSN respectively, supported the growth of the ΔrpfC strain to a maximum OD600 of 1.2. In XYSA medium, the ΔrpfC strain produced three DSF-family signals, DSF (0.9 μM or 2.0 × 10−3 μmol/109 cells), BDSF (2.8 μM or 6.2 × 10−3 μmol/109 cells) and IDSF (0.08 μM or 0.18 × 10−3 μmol/109 cells) (Fig. 3a). However, the ΔrpfC strain grown in XSYN medium produced only two DSF-family signals, DSF at a reduced level (0.22 μM or 0.48 × 10−3 μmol/109 cells), and BDSF (2.9 μM or 6.4 × 10−3 μmol/109 cells) (Fig. 3a). These results suggest that BCAAs are important for both DSF and IDSF biosynthesis.

Bottom Line: Furthermore, our biochemical analyses show that the key DSF synthase RpfF has both thioesterase and dehydratase activities, and uses 3-hydroxydedecanoyl-ACP as a substrate to produce BDSF.Finally, our results show that the classic fatty acid synthesis elongation cycle is required for the biosynthesis of DSF-family signals.Taken all together, these findings establish a general biosynthetic pathway for the DSF-family quorum sensing signals.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Microbial Metabolism, School of Life Sciences &Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, China.

ABSTRACT
Members of the diffusible signal factor (DSF) family are a novel class of quorum sensing (QS) signals in diverse Gram-negative bacteria. Although previous studies have identified RpfF as a key enzyme for the biosynthesis of DSF family signals, many questions in their biosynthesis remain to be addressed. In this study with the phytopathogen Xanthomonas campestris pv. campestris (Xcc), we show that Xcc produces four DSF-family signals (DSF, BDSF, CDSF and IDSF) during cell culture, and that IDSF is a new functional signal characterized as cis-10-methyl-2-dodecenoic acid. Using a range of defined media, we further demonstrate that Xcc mainly produces BDSF in the presence of carbohydrates; leucine and valine are the primary precursor for DSF biosynthesis; isoleucine is the primary precursor for IDSF biosynthesis. Furthermore, our biochemical analyses show that the key DSF synthase RpfF has both thioesterase and dehydratase activities, and uses 3-hydroxydedecanoyl-ACP as a substrate to produce BDSF. Finally, our results show that the classic fatty acid synthesis elongation cycle is required for the biosynthesis of DSF-family signals. Taken all together, these findings establish a general biosynthetic pathway for the DSF-family quorum sensing signals.

No MeSH data available.


Related in: MedlinePlus