RESUMEN
The human innate immunity factor apolipoprotein L-I (APOL1) protects against infection by several protozoan parasites, including Trypanosoma brucei brucei Endocytosis and acidification of high-density lipoprotein-associated APOL1 in trypanosome endosomes leads to eventual lysis of the parasite due to increased plasma membrane cation permeability, followed by colloid-osmotic swelling. It was previously shown that recombinant APOL1 inserts into planar lipid bilayers at acidic pH to form pH-gated nonselective cation channels that are opened upon pH neutralization. This corresponds to the pH changes encountered during endocytic recycling, suggesting APOL1 forms a cytotoxic cation channel in the parasite plasma membrane. Currently, the mechanism and domains required for channel formation have yet to be elucidated, although a predicted helix-loop-helix (H-L-H) was suggested to form pores by virtue of its similarity to bacterial pore-forming colicins. Here, we compare recombinant human and baboon APOL1 orthologs, along with interspecies chimeras and individual amino acid substitutions, to identify regions required for channel formation and pH gating in planar lipid bilayers. We found that whereas neutralization of glutamates within the H-L-H may be important for pH-dependent channel formation, there was no evidence of H-L-H involvement in either pH gating or ion selectivity. In contrast, we found two residues in the C-terminal domain, tyrosine 351 and glutamate 355, that influence pH gating properties, as well as a single residue, aspartate 348, that determines both cation selectivity and pH gating. These data point to the predicted transmembrane region closest to the APOL1 C terminus as the pore-lining segment of this novel channel-forming protein.
Asunto(s)
Apolipoproteína L1/química , Inmunidad Innata , Animales , Apolipoproteína L1/genética , Apolipoproteína L1/inmunología , Secuencias Hélice-Asa-Hélice , Humanos , Concentración de Iones de Hidrógeno , Papio hamadryas , Trypanosoma brucei brucei/inmunologíaRESUMEN
Mycobacterium tuberculosis is a major human pathogen and the causative agent of tuberculosis disease. M. tuberculosis is able to persist in the face of host-derived antimicrobial molecules nitric oxide (NO) and copper (Cu). However, M. tuberculosis with defective proteasome activity is highly sensitive to NO and Cu, making the proteasome an attractive target for drug development. Previous work linked NO susceptibility with the accumulation of para-hydroxybenzaldehyde (pHBA) in M. tuberculosis mutants with defective proteasomal degradation. In this study, we found that pHBA accumulation was also responsible for Cu sensitivity in these strains. We showed that exogenous addition of pHBA to wild-type M. tuberculosis cultures sensitized bacteria to Cu to a degree similar to that of a proteasomal degradation mutant. We determined that pHBA reduced the production and function of critical Cu resistance proteins of the regulated in copper repressor (RicR) regulon. Furthermore, we extended these Cu-sensitizing effects to an aldehyde that M. tuberculosis may face within the macrophage. Collectively, this study is the first to mechanistically propose how aldehydes can render M. tuberculosis susceptible to an existing host defense and could support a broader role for aldehydes in controlling M. tuberculosis infections. IMPORTANCE M. tuberculosis is a leading cause of death by a single infectious agent, causing 1.5 million deaths annually. An effective vaccine for M. tuberculosis infections is currently lacking, and prior infection does not typically provide robust immunity to subsequent infections. Nonetheless, immunocompetent humans can control M. tuberculosis infections for decades. For these reasons, a clear understanding of how mammalian immunity inhibits mycobacterial growth is warranted. In this study, we show aldehydes can increase M. tuberculosis susceptibility to copper, an established antibacterial metal used by immune cells to control M. tuberculosis and other microbes. Given that activated macrophages produce increased amounts of aldehydes during infection, we propose host-derived aldehydes may help control bacterial infections, making aldehydes a previously unappreciated antimicrobial defense.
Asunto(s)
Mycobacterium tuberculosis , Tuberculosis , Animales , Humanos , Mycobacterium tuberculosis/metabolismo , Cobre/farmacología , Cobre/metabolismo , Aldehídos/metabolismo , Aldehídos/farmacología , Complejo de la Endopetidasa Proteasomal/metabolismo , Tuberculosis/microbiología , Antibacterianos/farmacología , Mamíferos/metabolismoRESUMEN
Most ant species have two distinct female castes-queens and workers-yet the developmental and genetic mechanisms that produce these alternative phenotypes remain poorly understood. Working with a clonal ant, we discovered a variant strain that expresses queen-like traits in individuals that would normally become workers. The variants show changes in morphology, behavior, and fitness that cause them to rely on workers in wild-type (WT) colonies for survival. Overall, they resemble the queens of many obligately parasitic ants that have evolutionarily lost the worker caste and live inside colonies of closely related hosts. The prevailing theory for the evolution of these workerless social parasites is that they evolve from reproductively isolated populations of facultative intermediates that acquire parasitic phenotypes in a stepwise fashion. However, empirical evidence for such facultative ancestors remains weak, and it is unclear how reproductive isolation could gradually arise in sympatry. In contrast, we isolated these variants just a few generations after they arose within their WT parent colony, implying that the complex phenotype reported here was induced in a single genetic step. This suggests that a single genetic module can decouple the coordinated mechanisms of caste development, allowing an obligately parasitic variant to arise directly from a free-living ancestor. Consistent with this hypothesis, the variants have lost one of the two alleles of a putative supergene that is heterozygous in WTs. These findings provide a plausible explanation for the evolution of ant social parasites and implicate new candidate molecular mechanisms for ant caste differentiation.