CcpA Regulates Arginine Biosynthesis in <em>Staphylococcus aureus</em> through Repression of Proline Catabolism

<div><p><em>Staphylococcus aureus</em> is a leading cause of community-associated and nosocomial infections. Imperative to the success of <em>S. aureus</em> is the ability to adapt and utilize nutrients that are readily available. Genomic sequencing suggests that <em>S. aureus</em> has the genes required for synthesis of all twenty amino acids. However, <em>in vitro</em> experimentation demonstrates that staphylococci have multiple amino acid auxotrophies, including arginine. Although <em>S. aureus</em> possesses the highly conserved anabolic pathway that synthesizes arginine via glutamate, we demonstrate here that inactivation of <em>ccpA</em> facilitates the synthesis of arginine via the urea cycle utilizing proline as a substrate. Mutations within <em>putA</em>, <em>rocD</em>, <em>arcB1</em>, <em>argG</em> and <em>argH</em> abolished the ability of <em>S. aureus</em> JE2 <em>ccpA::tetL</em> to grow in the absence of arginine, whereas an interruption in <em>argJBCF</em>, <em>arcB2</em>, or <em>proC</em> had no effect. Furthermore, nuclear magnetic resonance demonstrated that JE2 <em>ccpA::ermB</em> produced ¹³C₅ labeled arginine when grown with ¹³C₅ proline. Taken together, these data support the conclusion that <em>S. aureus</em> synthesizes arginine from proline during growth on secondary carbon sources. Furthermore, although highly conserved in all sequenced <em>S. aureus</em> genomes, the arginine anabolic pathway (ArgJBCDFGH) is not functional under <em>in vitro</em> growth conditions. Finally, a mutation in <em>argH</em> attenuated virulence in a mouse kidney abscess model in comparison to wild type JE2 demonstrating the importance of arginine biosynthesis <em>in vivo</em> via the urea cycle. However, mutations in <em>argB</em>, <em>argF</em>, and <em>putA</em> did not attenuate virulence suggesting both the glutamate and proline pathways are active and they, or their pathway intermediates, can complement each other <em>in vivo</em>.</p>