Posttranslational Maturation of the Invasion Acyl Carrier Protein of Salmonella enterica Serovar Typhimurium Requires an Essential Phosphopantetheinyl Transferase of the Fatty Acid Biosynthesis Pathway

Abstract

c Salmonella pathogenicity island 1 (SPI-1) carries genes required for the formation of a type 3 secretion system, which is necessary for the invasion process of Salmonella. Among the proteins encoded by SPI-1 is IacP, a homolog of acyl carrier proteins. Acyl carrier proteins are mainly involved in fatty acid biosynthesis, and they require posttranslational maturation by addition of a 4=-phosphopantetheine prosthetic group to be functional. In this study, we analyzed IacP maturation in vivo. By performing matrix-assisted laser desorption ionization-time-of-flight (MALDI-TOF) mass spectrometry analysis of intact purified proteins, we showed that IacP from Salmonella enterica serovar Typhimurium was matured by addition of 4=-phosphopantetheine to the conserved serine 38 residue. Therefore, we searched for the phosphopantetheinyl transferases in charge of IacP maturation. A bacterial two-hybrid approach revealed that IacP interacted with AcpS, an enzyme normally required for the maturation of the canonical acyl carrier protein (ACP), which is involved in fatty acid biosynthesis. The creation of a conditional acpS mutant then demonstrated that AcpS was necessary for the maturation of IacP. However, although IacP was similar to ACP and matured by using the same enzyme, IacP could not replace the essential function of ACP in fatty acid synthesis. Hence, the demonstration that IacP is matured by AcpS establishes a cross-connection between virulence and fatty acid biosynthesis pathways.

Acylation of the Type 3 Secretion System Translocon Using a Dedicated Acyl Carrier Protein

“…The S . Typhimurium IacP_TAP strain was used, in which the TAP tag sequence had been introduced in frame with the 3’ end of iacP at the chromosomal locus ( S1 Fig ) [ 18 ]. The TAP tag includes two IgG binding domains of Staphylococcus aureus protein A (ProtA) and a calmodulin binding peptide (CBP) separated by a TEV protease cleavage.…”

“…IacP was found to interact with three proteins: i/ FabB, which is the ß - ketoacyl-ACP synthase I involved in saturated and unsaturated fatty acid biosynthesis, ii/ IscS, which is a cysteine desulfurase crucial for iron-sulfur cluster biogenesis and repair, and iii/ SipB, the major translocator of SPI-1 T3SS. The 4’-PP modification of IacP was required for the interaction of IacP with these three proteins since none of them were retrieved when the TAP experiment was performed using a strain producing the IacP S38T _TAP protein that had lost capacity to be modified by a 4’-PP group [ 18 ] ( Fig 1B and S1 Fig ).…”

“…We previously showed that IacP is modified by a 4’-PP group on the conserved serine 38 residue, as expected for an acyl carrier protein [ 18 ]. In this study, we investigated the role of iacP to understand its association to SPI-1.…”

Salmonella—how a metabolic generalist adopts an intracellular lifestyle during infection

“…Various metabolic pathways which have an impact on the SPI1 activity of Salmonella enterica (Table 3 ). One example is the interaction between the invasion acyl carrier protein (IacP; Viala et al, 2013 ) and secretion of SPI1 effector proteins into the host cell to achieve rearrangement of the host cytoskeleton and engulfment of the bacterium (reviewed in Cossart and Sansonetti, 2004 ).…”

Meeting report: Adaptation and communication of bacterial pathogens

“…Its corresponding gene as well as those encoding the T3SS are all localized within the SPI-1 gene cluster. 33 Her work showed that the SPI-1 T3SS is posttranslationally modified according to a process that involves IacP, leading to the optimization of the poreforming activity of the machinery.…”