ATF6 enables pathogen infection in ticks by inducing <i>stomatin</i> and altering cholesterol dynamics

How tick-borne pathogens interact with their hosts has been primarily studied in vertebrates where disease is observed. Comparatively less is known about pathogen interactions within the tick. Here, we report that <i>Ixodes scapularis</i> ticks infected with either <i>Anaplasma phagocytophilum</i> (causative agent of anaplasmosis) or <i>Borrelia burgdorferi</i> (causative agent of Lyme disease) show activation of the ATF6 branch of the unfolded protein response (UPR). Disabling ATF6 functionally restricts pathogen survival in ticks. When stimulated, ATF6 functions as a transcription factor, but is the least understood out of the three UPR pathways. To interrogate the <i>Ixodes</i> ATF6 transcriptional network, we developed a custom R script to query tick promoter sequences. This revealed <i>stomatin</i> as a potential gene target, which has roles in lipid homeostasis and vesical transport. <i>Ixodes stomatin</i> was experimentally validated as a bona fide ATF6-regulated gene through luciferase reporter assays, pharmacological activators, RNA interference transcriptional repression, and immunofluorescence microscopy. Silencing <i>stomatin</i> decreased <i>A. phagocytophilum</i> colonization in <i>Ixodes</i> and disrupted cholesterol dynamics in tick cells. Furthermore, blocking <i>stomatin</i> restricted cholesterol availability to the bacterium, thereby inhibiting growth and survival. Taken together, we have identified the <i>Ixodes</i> ATF6 pathway as a contributor to vector competence through Stomatin-regulated cholesterol homeostasis. Moreover, our custom, web-based transcription factor binding site search tool "ArthroQuest" revealed that the ATF6-regulated nature of <i>stomatin</i> is unique to blood-feeding arthropods. Collectively, these findings highlight the importance of studying fundamental processes in nonmodel organisms.