UCLA

Understanding host-pathogen interactions between microbes and the innate immune system will provide insight into the host defense pathways and microbial virulence factors. The macrophage (MΦ) is a sentinel of the innate immune system and serves as the first line of defense against microbial infection. The MΦ provides protection to the host by i) rapidly recognizing harmful pathogens, ii) internalizing pathogens to contain the infection and iii) clearing the pathogen from the host before the onset of disease. Here, we study effector pathways of the MΦ that combat invading pathogens in the host and determine their effects on the invading pathogen.

MΦ are a phenotypically heterogeneous immune subset that provides different functions in host defense. Previous studies in our laboratory have found that interleukin-15 (IL-15) induces a MΦ differentiation program in primary human monocytes (IL-15 MΦ) that express the vitamin D metabolism pathway. Vitamin D supplementation to these specialized immune subsets exhibited an antimicrobial response against mycobacteria in vitro. However, clinical trials that supplemented tuberculosis (TB) patients with vitamin D as an adjuvant have largely been unsuccessful in vivo. Therefore, we investigate whether vitamin D status prior to the onset of microbial infection would contribute to host defense. Our data demonstrates that vitamin D status during IL-15 MΦ differentiation bestows the capacity to mount an antimicrobial response against Mycobacterium leprae. These data suggest that future clinical trials that assess the relationship between vitamin D supplementation to mycobacterial infection will determine if vitamin D can provide prophylactic effects that are therapeutically beneficial.

Similarly to vitamin D, the bioactive form of vitamin A, all-trans retinoic acid (ATRA), triggers an antimicrobial responses against M. tuberculosis in vitro. However, high ATRA levels in humans’ results in severe or even fatal side effects in vivo. Therefore, we investigate how the immune system regulates ATRA production at the site of TB disease. Our data demonstrates that dendritic cells express the vitamin A metabolism pathway, which converts the circulatory form of vitamin A, retinol, into ATRA. The dendritic cells subsequently release ATRA and induce vitamin A-dependent antimicrobial responses in neighboring monocytes and MΦ. Interestingly, this immune model has provided insight into the site of TB disease by showing that the dendritic cell-mediated retinol metabolism pathway is significantly diminished in the lung of active TB patients relative to normal lung. These data demonstrate a novel transcellular effector pathway between dendritic cells and MΦ that contributes to the host defense against microbial infection.

Toxoplasma gondii is capable of infecting any nucleated cell in vitro, but MΦ are the first immune subset infected by T. gondii in mice in vivo. It is well known that both interferon-gamma-induced responses and MΦ functions are critical to controlling T. gondii infections in mice in vivo. Interferon-gamma induces the expression of two different families of immune loading proteins called the immunity-related GTPases (IRGs) and guanylate binding proteins (GBPs) that function to clear the parasite from the MΦ. However, T. gondii is equipped with secretory organelles called the rhoptries that inject ROP proteins into MΦ to modulate host cell functions. We have found a novel rhoptry pseudokinase effector, ROP54. Disruption of ROP54 demonstrates a 100-fold decrease in virulence in mice in vivo and increased GBP2 protein loading onto the parasite containing vacuole in vitro. Immunoprecipitation of ROP54 demonstrates that none of the known ROP effector proteins formed a complex with the pseudokinase, which suggests it may be a divergent effector protein. Collectively these data show that ROP54 is a novel virulence factor that evades a IFN-gamma-mediated immune response in MΦ and may be a potential drug target for the development of novel therapeutics.

The data presented in this thesis evaluates the efficiency of immune pathways against disease causing pathogens. The host-pathogen interaction from these models provides insight into microbial infection, which may help in the development of novel therapeutics and identification of drug targets. These data contributes to the importance of micronutrient supplementation and how it can help contain the spread of mycobacterial-related diseases. Additionally we found a novel ROP effector protein that may be a potential drug target in toxoplasma infection.