Protein synthesis in Brucella abortus induced during macrophage infection

Abstract

Brucella abortus is a facultative, intracellular, pathogenic bacterium that replicates within macrophages and resists macrophage microbicidal mechanisms. To study gene expression and to elucidate the defense mechanisms used by B. abortus to resist destruction within macrophages, protein synthesis by B. abortus was examined by pulse-labeling techniques during intracellular growth within J774A.1, a macrophage-like cell line. Prominent changes observed include increased synthesis of Brucella proteins with estimated molecular masses of 62, 28, 24, and 17 kDa. The 62-kDa protein was identified by immunoprecipitation analysis as Hsp62, a GroEL homolog. A protein of 60 kDa was expressed during acid shock and may represent a modified form of Hsp62. The 28-and 17-kDa proteins have not been observed under any in vitro stress condition and presumably represent macrophage-specific induction. The 24-kDa protein comigrates with an unidentified protein induced by acid shock, designated Asp24. Expression of Asp24 is optimal at pH values below 4.0 and within the first 3 h following a shift from pH 7.3 to 3.8. This corresponds directly with a period of optimal bacterial survival at a reduced pH and suggests an active role for this protein in resistance to such environments. The identification of these gene products and the mechanisms controlling their expression is an important step in understanding the resistance of Brucella spp. to intracellular destruction within macrophages.

Participation of the molecular chaperone DnaK in intracellular growth of Brucella suis within U937‐derived phagocytes

“…The authors describe an increase in the synthesis of proteins having apparent molecular masses of 70, 62, 18, and 10 kDa under heat-shock conditions, among which only Hsp 62 has been identified, being the GroEL homologue. More recently, these authors speculated that the 70 kDa protein was DnaK (Lin and Ficht, 1995). Our results confirmed the heat-shock induction of proteins with apparent molecular masses of 70, 62, and 10 kDa in B. suis, but we also found induction of proteins of 36, 20.5, 16, and 14.5 kDa.…”

“…In agreement with these findings, our experimental results showed the induction of DnaK at low pH, and that a DnaK-producing strain of B. suis was significantly more resistant to acid pH than a dnaK null mutant. The protein profile of 6. abortus under low pH conditions has been published recently (Lin and Ficht, 1995), showing the induction of the two major proteins Asp-24 and Asp-60, of which the latter may represent a modified form of the GroEL homologue.…”

“…In addition to performing in vitro experiments, Lin and Ficht (1995) also described the protein profile of B. abortus after infection of J774 murine macrophages. Strengthening our findings, they observe increased synthesis of seven proteins among which is the 70 kDa protein, presumably DnaK.…”

Cold shock protein A plays an important role in the stress adaptation and virulence ofBrucella melitensis

“…Moreover, the cspA gene was also found to be induced by oxidative stress. Although the functional contributions of CspA to stress tolerance in Brucella are not yet clear, research has shown that Brucella that are grown in macrophages undergo a significant change in protein expression (Lin & Ficht, 1995;Rafie-Kolpin et al, 1996). Because the RNA chaperone Hfq acts as a global regulator in Brucella (Caswell et al, 2012) and CspA is also an RNA chaperone (Jiang et al, 1997) that is linked to the regulation of the expression of multiple genes in E. coli (Bae et al, 1997;Phadtare & Inouye, 2004), an intriguing possibility is that CspA may participate in the regulation of some or all of these proteins involved in stress tolerance.…”

Usefulness of a quantitative real-time PCR assay using serum samples to discriminate between inactive, serologically positive and active human brucellosis

“…Brucellosis remains the most common zoonotic infection worldwide, representing a major form of disease in humans [1]. Brucella is an intracellular pathogen and is able to elude fusion by phagolysosomes, thereby enabling it to survive and even multiply within cells of the mononuclear phagocyte system [2]. This intracellular survival determines the characteristic clinical profile of the disease, i.e.…”