Adeno-associated virus (AAV)-mediated genetic targeting of microglia remains a challenge. Overcoming this hurdle is essential for gene editing in the central nervous system (CNS). Here, we characterized the minimal/native promoter of the HEXB gene, which is known to be specifically and stably expressed in the microglia during homeostatic and pathological conditions. Dual reporter and serial deletion assays identified the critical role of the natural 5’ untranslated region (−97 bp related to the first ATG) in driving transcriptional activity of the mouse Hexb gene. The native promoter region of mouse, human, and monkey HEXB are located at −135, −134, and −170 bp to the first ATG, respectively. These promoters were highly active and specific in microglia with strong cross-species transcriptional activities, but did not exhibit activity in primary astrocytes. In addition, we identified a 135 bp promoter of CD68 gene that was highly active in microglia but not in astrocytes. Considering that HEXB is specifically expressed in microglia, these data suggest that the newly characterized microglia-specific HEXB minimal/native promoter can be an ideal candidate for microglia-targeting AAV gene therapy in the CNS.

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Adeno-associated virus (AAV)-mediated genetic targeting of microglia remains challenging. Overcoming this hurdle is essential for gene editing in the central nervous system (CNS). Here, we characterized the minimal/native promoter of the HexB gene, which is known to be specifically and stably expressed in the microglia during homeostatic and pathological conditions. Dual reporter and serial deletion assays identified the critical role of the natural 5’ untranslated region (−97 bp related to the first ATG) in driving transcriptional activity of the mouse HexB gene. The native promoter region of mouse, human and monkey HexB located at -135, -134 and -170 bp to the first ATG, respectively. These promoters were highly active and specific in microglia with strong cross-species transcriptional activities, but had no activities in primary astrocytes. In addition, we identified a 135 bp promoter of CD68 gene was also highly active in microglia but not in astrocytes. Considering that HexB is specifically expressed in microglia, not in monocytes/macrophages or other neuronal cells, these data suggest that the newly characterized 134 bp microglia-specific HexB promoter can be an ideal candidate for microglia-targeting AAV gene therapy, which could be developed for HIV eradication in the brain wherein microglia harbor the main HIV reservoirs in the CNS.SummaryIt is hard to overstate the importance of gene therapy that can remove viral genes from human cells. A cure for HIV would mean a lifetime free of treatment for patients who now must maintain a strict regimen of ART indefinitely. In order to develop a cure using AAV delivery, payload DNA must meet the AAV vector size limitations, and the payload genes must be expressed appropriately. Previous studies have identified microglia-specific HexB gene that shows stable expression during neural homeostasis and pathogenesis. Our study identified the essential HexB gene promoter (134 bp) as a strong candidate for AAV gene therapy to specifically target the brain microglia, the main cellular reservoirs of HIV in the central nervous system. Our studies continue to move us closer to identifying target-specific gene therapy for NeuroAIDS.

Strauss, 1967). Seventeen individual interviews, and one focus group of seven, were conducted to gather data. Constant comparison of the findings and analysis was conducted until theoretical saturation was reached (Glaser & Strauss, 1967). Findings were presented to the individuals involved in the focus group, and to experts in the field, for validation (Lincoln & Guba, 1985). Results Analysis of the findings revealed that nurses felt they had become prepared through a feeling of 'fitting' with their workplace, colleagues and patients, through a shared ideal way to care for people at end of life. Comparison of findings with the literature, identified that the Person/Environment Fit (PEF) theory (Edwards & Billsberry, 2010) could be used to conceptualise these findings. According to PEF theory, people choose the environment that they feel fits their vocational ideals, as well as where they fit in with the organisation's philosophy, their work group, job role and other individuals in the workplace (Edwards & Billsberry, 2010). However, as hospices evolve rapidly to ensure accessibility to more patients and families than ever before (Mathew, Gray & Thomas, 2018), participants commented on the changing work environment. People who felt a strong feeling of 'fit' on joining the hospice, do not always feel that fit is still there. Conclusion As hospices continue to transform from a 'Rolls Royce' service for the minority, into a more modest style for all (Mathew, Gray & Thomas, 2018), the findings of this study demonstrate that, during transformation, hospices may need to take measures to maintain an 'ideal' way of caring, that people feel fits their vocational aspirations.