Advisory Committee Chair
Advisory Committee Members
Date of Award
Degree Name by School
Doctor of Philosophy (PhD) School of Optometry
X-genetics aims to replace visible light stimulation in optogenetics with X-radiation, offering improved bone penetration, minimal scattering, and enhanced targeting capabilities. This eliminates the need for surgical optic fiber implantation, reducing invasiveness and minimizing off-target effects. Discovering a receptor protein sensitive to X-rays or their byproducts is pivotal for advancing X-genetics.Two genetically encoded X-ray receptors (GEXRs) were investigated for X-genetics application: rhodopsin, historically associated with X-ray responses, and LITE-1, an unexpected candidate. Rhodopsin, when expressed in human embryonic kidney (HEK293) cells, demonstrated robust cAMP decreases upon visible light stimulation but remained unresponsive to moderate and high-dose X-rays, suggesting it may not be suitable as a GEXR. LITE-1, initially explored for UV sensitivity, surprisingly exhibited X-ray sensitivity in wild type C. elegans, as indicated by a short latency X-ray avoidance response absent in lite-1 mutants. Ectopic LITE-1 expression in C. elegans muscle cells mediated X-ray-induced muscle contraction and paralysis responses, demonstrating the receptor’s capability to confer X-ray sensitivity to otherwise insensitive cells. To assess LITE-1's potential as an X-ray receptor in mammalian systems, lite-1::1D4 was cloned into a mammalian expression vector with which HEK293 cells were transfected. While robustLITE-1 expression localized to the plasma membrane was observed, cAMP luminescence-based GsO assays failed to detect g-protein activation in response to UV or X-ray stimulation. Immunolabeling of membrane-intact cells failed to label LITE-1, suggesting improper expression in HEK293 cells. These findings help identify GEXRs for X-genetics. Rhodopsin's inability to confer X-ray sensitivity to HEK293 cells challenges its role as an X-ray receptor. On the other hand, LITE-1 successfully imparts X-ray sensitivity in C. elegans but fails to do so in HEK293 cells, likely due to membrane trafficking issues. These results eliminate one potential GEXR while introducing a new GEXR with promise in C. elegans. Further research in mammalian neurons is necessary to fully explore LITE-1's potential for X-genetics in mammals.
Video_1_LITE-1 mediates behavioral responses to X-rays in Caenorhabditis elegans.MP4 (16997 kB)
Video_2_LITE-1 mediates behavioral responses to X-rays in Caenorhabditis elegans.MP4 (17175 kB)
Video_3_LITE-1 mediates behavioral responses to X-rays in Caenorhabditis elegans.MP4 (28549 kB)
Video_4_LITE-1 mediates behavioral responses to X-rays in Caenorhabditis elegans.MP4 (19423 kB)
Video_5_LITE-1 mediates behavioral responses to X-rays in Caenorhabditis elegans.MP4 (29615 kB)
Video_6_LITE-1 mediates behavioral responses to X-rays in Caenorhabditis elegans.MP4 (29630 kB)
Video_7_LITE-1 mediates behavioral responses to X-rays in Caenorhabditis elegans.MP4 (16927 kB)
Cannon, Kelli, "Towards Minimally Invasive Genetically Targeted Control Of Neural Activity Using X-Rays" (2023). All ETDs from UAB. 3484.