Sara Castaño-cerezo | Universidad de Murcia (original) (raw)

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Papers by Sara Castaño-cerezo

Molecular Systems Biology, 2014

Although protein acetylation is widely observed, it has been associated with few specific regulat... more Although protein acetylation is widely observed, it has been associated with few specific regulatory functions making it poorly understood. To interrogate its functionality, we analyzed the acetylome in Escherichia coli knockout mutants of cobB, the only known sirtuin-like deacetylase, and patZ, the best-known protein acetyltransferase. For four growth conditions, more than 2,000 unique acetylated peptides, belonging to 809 proteins, were identified and differentially quantified. Nearly 65% of these proteins are related to metabolism. The global activity of CobB contributes to the deacetylation of a large number of substrates and has a major impact on physiology. Apart from the regulation of acetyl-CoA synthetase, we found that CobB-controlled acetylation of isocitrate lyase contributes to the fine-tuning of the glyoxylate shunt. Acetylation of the transcription factor RcsB prevents DNA binding, activating flagella biosynthesis and motility, and increases acid stress susceptibility. Surprisingly, deletion of patZ increased acetylation in acetate cultures, which suggests that it regulates the levels of acetylating agents. The results presented offer new insights into functional roles of protein acetylation in metabolic fitness and global cell regulation.

Applied microbiology and biotechnology, Jan 20, 2014

Acetate production is one of the most striking differences between Escherichia coli K12 and BL21 ... more Acetate production is one of the most striking differences between Escherichia coli K12 and BL21 strains. Transcription of acetate metabolism genes is regulated. Additionally, acetyl-CoA synthetase, which activates acetate to acetyl-CoA, is regulated by post-translational acetylation. The aim of this study was to understand the contribution of reversible protein lysine acetylation to the regulation of acetate metabolism in E. coli BL21. The phenotypic differences between both strains were especially important in the presence of acetate. The high expression of acetyl-CoA synthetase (acs) in glucose exponential phase in BL21 allows the simultaneous consumption of acetate and glucose. Lack of catabolite repression also affected its post-translational regulator, the protein acetyltransferase (patZ). The effect of the deletion of cobB (encoding a sirtuin-like protein deacetylase) and patZ genes depended on the genetic background. The deletion of cobB in both strains increased acetate pro...

New Biotechnology, 2014

Post-translational modification of proteins is a reversible mechanism of cellular adaptation to c... more Post-translational modification of proteins is a reversible mechanism of cellular adaptation to changing environmental conditions. In eukaryotes, the physiological relevance of N-ɛ-lysine protein acetylation is well demonstrated. In recent times, important roles in the regulation of metabolic processes in bacteria are being uncovered, adding complexity to cellular regulatory networks. The aim of this mini-review is to sum up the current state-of-the-art in the regulation of bacterial physiology by protein acetylation. Current knowledge on the molecular biology aspects of known bacterial protein acetyltransferases and deacetylases will be summarized. Protein acetylation in Escherichia coli, Salmonella enterica, Bacillus subtilis, Rhodopseudomonas palustris and Mycobacterium tuberculosis, will be explained in the light of their physiological relevance. Progress in the elucidation of bacterial acetylomes and the emerging understanding of chemical acylation mechanisms will be discussed together with their regulatory and evolutionary implications. Fundamental molecular studies detailing this recently discovered regulatory mechanism pave the way for their prospective application for the construction of synthetic regulation networks.

Molecular Microbiology, 2011

Lysine acetylation is a well-established post-translational modification widely conserved and dis... more Lysine acetylation is a well-established post-translational modification widely conserved and distributed in bacteria. Although multiple regulatory roles have been proved, little is known about its regulation. Here, we present evidence that the transcription of the Gcn5-like acetyltransferase YfiQ of Escherichia coli (proposed name: PatZ) is regulated by cAMP-CRP and its implications on acetate metabolism regulation. The acetate scavenging acetyl-CoA synthetase (Acs) is regulated at the transcriptional and post-translational levels. Post-translational regulation depends on a protein acetyltransferase (yfiQ) and an NAD(+) -dependent deacetylase (cobB). We have studied their expression under different environmental conditions. cobB is constitutively expressed from a promoter located upstream nagK. The expression of yfiQ occurs from its own promoter; it is upregulated in the stationary phase and in the presence of non-PTS carbon sources and is positively regulated by cAMP-CRP. Two putative CRP binding sites are necessary for its full activity. Gene deletion revealed that cobB is essential for growth on acetate, yfiQ deletion restoring growth of the cobB mutant. The fine tuning of metabolic enzymes results from the integration of multiple mechanisms, and redundant systems may exist. Despite the existence of divergent catabolite repression systems, this may be a conserved strategy common to both Gram-positive and -negative bacteria.

Microbial Cell Factories, 2009

Background: Acetate metabolism in Escherichia coli plays an important role in the control of the ... more Background: Acetate metabolism in Escherichia coli plays an important role in the control of the central metabolism and in bioprocess performance. The main problems related to the use of E. coli as cellular factory are i) the deficient utilization of carbon source due to the excretion of acetate during aerobic growth, ii) the inhibition of cellular growth and protein production by acetate and iii) the need for cofactor recycling (namely redox coenzymes and free CoASH) to sustain balanced growth and cellular homeostasis.

Impairment of acetate production in Escherichia coli is crucial for the performance of many biote... more Impairment of acetate production in Escherichia coli is crucial for the performance of many biotechnological processes. Aerobic production of acetate (or acetate overflow) results from changes in the expression of central metabolism genes. Acetyl-CoA synthetase scavenges extracellular acetate in glucose-limited cultures. Once converted to acetyl-CoA, it can be catabolized by the tricarboxylic acid cycle or the glyoxylate pathway. In this work, we assessed the significance of these pathways on acetate overflow during glucose excess and limitation. Gene expression, enzyme activities, and metabolic fluxes were studied in E. coli knock-out mutants related to the glyoxylate pathway operon and its regulators. The relevance of post-translational regulation by AceK-mediated phosphorylation of isocitrate dehydrogenase for pathway functionality was underlined. In chemostat cultures performed at increasing dilution rates, acetate overflow occurs when growing over a threshold glucose uptake rate. This threshold was not affected in a glyoxylate-pathway-deficient strain (lacking isocitrate lyase, the first enzyme of the pathway), indicating that it is not relevant for acetate overflow. In carbon-limited chemostat cultures, gluconeogenesis (maeB, sfcA, and pck), the glyoxylate operon and, especially, acetyl-CoA synthetase are upregulated. A mutant in acs (encoding acetyl-CoA synthetase) produced acetate at all dilution rates. This work demonstrates that, in E. coli, acetate production occurs at all dilution rates and that overflow is the result of unbalanced synthesis and scavenging activities. The over-expression of acetyl-CoA synthetase by cAMP-CRP-dependent induction limits this phenomenon in cultures consuming glucose at low rate, ensuring the recycling of the acetyl-CoA and acetyl-phosphate pools, although establishing an energy-dissipating substrate cycle.

Molecular Systems Biology, 2014

Although protein acetylation is widely observed, it has been associated with few specific regulat... more Although protein acetylation is widely observed, it has been associated with few specific regulatory functions making it poorly understood. To interrogate its functionality, we analyzed the acetylome in Escherichia coli knockout mutants of cobB, the only known sirtuin-like deacetylase, and patZ, the best-known protein acetyltransferase. For four growth conditions, more than 2,000 unique acetylated peptides, belonging to 809 proteins, were identified and differentially quantified. Nearly 65% of these proteins are related to metabolism. The global activity of CobB contributes to the deacetylation of a large number of substrates and has a major impact on physiology. Apart from the regulation of acetyl-CoA synthetase, we found that CobB-controlled acetylation of isocitrate lyase contributes to the fine-tuning of the glyoxylate shunt. Acetylation of the transcription factor RcsB prevents DNA binding, activating flagella biosynthesis and motility, and increases acid stress susceptibility. Surprisingly, deletion of patZ increased acetylation in acetate cultures, which suggests that it regulates the levels of acetylating agents. The results presented offer new insights into functional roles of protein acetylation in metabolic fitness and global cell regulation.

Applied microbiology and biotechnology, Jan 20, 2014

Acetate production is one of the most striking differences between Escherichia coli K12 and BL21 ... more Acetate production is one of the most striking differences between Escherichia coli K12 and BL21 strains. Transcription of acetate metabolism genes is regulated. Additionally, acetyl-CoA synthetase, which activates acetate to acetyl-CoA, is regulated by post-translational acetylation. The aim of this study was to understand the contribution of reversible protein lysine acetylation to the regulation of acetate metabolism in E. coli BL21. The phenotypic differences between both strains were especially important in the presence of acetate. The high expression of acetyl-CoA synthetase (acs) in glucose exponential phase in BL21 allows the simultaneous consumption of acetate and glucose. Lack of catabolite repression also affected its post-translational regulator, the protein acetyltransferase (patZ). The effect of the deletion of cobB (encoding a sirtuin-like protein deacetylase) and patZ genes depended on the genetic background. The deletion of cobB in both strains increased acetate pro...

New Biotechnology, 2014

Post-translational modification of proteins is a reversible mechanism of cellular adaptation to c... more Post-translational modification of proteins is a reversible mechanism of cellular adaptation to changing environmental conditions. In eukaryotes, the physiological relevance of N-ɛ-lysine protein acetylation is well demonstrated. In recent times, important roles in the regulation of metabolic processes in bacteria are being uncovered, adding complexity to cellular regulatory networks. The aim of this mini-review is to sum up the current state-of-the-art in the regulation of bacterial physiology by protein acetylation. Current knowledge on the molecular biology aspects of known bacterial protein acetyltransferases and deacetylases will be summarized. Protein acetylation in Escherichia coli, Salmonella enterica, Bacillus subtilis, Rhodopseudomonas palustris and Mycobacterium tuberculosis, will be explained in the light of their physiological relevance. Progress in the elucidation of bacterial acetylomes and the emerging understanding of chemical acylation mechanisms will be discussed together with their regulatory and evolutionary implications. Fundamental molecular studies detailing this recently discovered regulatory mechanism pave the way for their prospective application for the construction of synthetic regulation networks.

Molecular Microbiology, 2011

Lysine acetylation is a well-established post-translational modification widely conserved and dis... more Lysine acetylation is a well-established post-translational modification widely conserved and distributed in bacteria. Although multiple regulatory roles have been proved, little is known about its regulation. Here, we present evidence that the transcription of the Gcn5-like acetyltransferase YfiQ of Escherichia coli (proposed name: PatZ) is regulated by cAMP-CRP and its implications on acetate metabolism regulation. The acetate scavenging acetyl-CoA synthetase (Acs) is regulated at the transcriptional and post-translational levels. Post-translational regulation depends on a protein acetyltransferase (yfiQ) and an NAD(+) -dependent deacetylase (cobB). We have studied their expression under different environmental conditions. cobB is constitutively expressed from a promoter located upstream nagK. The expression of yfiQ occurs from its own promoter; it is upregulated in the stationary phase and in the presence of non-PTS carbon sources and is positively regulated by cAMP-CRP. Two putative CRP binding sites are necessary for its full activity. Gene deletion revealed that cobB is essential for growth on acetate, yfiQ deletion restoring growth of the cobB mutant. The fine tuning of metabolic enzymes results from the integration of multiple mechanisms, and redundant systems may exist. Despite the existence of divergent catabolite repression systems, this may be a conserved strategy common to both Gram-positive and -negative bacteria.

Microbial Cell Factories, 2009

Background: Acetate metabolism in Escherichia coli plays an important role in the control of the ... more Background: Acetate metabolism in Escherichia coli plays an important role in the control of the central metabolism and in bioprocess performance. The main problems related to the use of E. coli as cellular factory are i) the deficient utilization of carbon source due to the excretion of acetate during aerobic growth, ii) the inhibition of cellular growth and protein production by acetate and iii) the need for cofactor recycling (namely redox coenzymes and free CoASH) to sustain balanced growth and cellular homeostasis.

Impairment of acetate production in Escherichia coli is crucial for the performance of many biote... more Impairment of acetate production in Escherichia coli is crucial for the performance of many biotechnological processes. Aerobic production of acetate (or acetate overflow) results from changes in the expression of central metabolism genes. Acetyl-CoA synthetase scavenges extracellular acetate in glucose-limited cultures. Once converted to acetyl-CoA, it can be catabolized by the tricarboxylic acid cycle or the glyoxylate pathway. In this work, we assessed the significance of these pathways on acetate overflow during glucose excess and limitation. Gene expression, enzyme activities, and metabolic fluxes were studied in E. coli knock-out mutants related to the glyoxylate pathway operon and its regulators. The relevance of post-translational regulation by AceK-mediated phosphorylation of isocitrate dehydrogenase for pathway functionality was underlined. In chemostat cultures performed at increasing dilution rates, acetate overflow occurs when growing over a threshold glucose uptake rate. This threshold was not affected in a glyoxylate-pathway-deficient strain (lacking isocitrate lyase, the first enzyme of the pathway), indicating that it is not relevant for acetate overflow. In carbon-limited chemostat cultures, gluconeogenesis (maeB, sfcA, and pck), the glyoxylate operon and, especially, acetyl-CoA synthetase are upregulated. A mutant in acs (encoding acetyl-CoA synthetase) produced acetate at all dilution rates. This work demonstrates that, in E. coli, acetate production occurs at all dilution rates and that overflow is the result of unbalanced synthesis and scavenging activities. The over-expression of acetyl-CoA synthetase by cAMP-CRP-dependent induction limits this phenomenon in cultures consuming glucose at low rate, ensuring the recycling of the acetyl-CoA and acetyl-phosphate pools, although establishing an energy-dissipating substrate cycle.