Using RNA-Seq to Identify Novel Pathomechanisms in Recessive X-Linked Ichthyosis
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The top layer of skin, the epidermis, is responsible for providing a protective barrier between an organism and its environment. The importance of the epidermis and its barrier functions are shown by the severity of diseases in which there is impaired barrier function, such as the ichthyoses. Recessive X-linked ichthyosis, caused by deletion or point mutations of the STS gene is one of the most common ichthyoses, but there is a paucity of data regarding this disease and its pathomechanisms. My hypothesis was that introducing knockdown of STS into immortalized N/TERT keratinocytes, which are then used to develop a 3D model of the skin, will replicate the disease process. Stable knockdowns of STS were generated in immortalized N/TERT keratinocytes. Knockdown of STS altered histology, lipid content, differentiation, desquamation, and transglutaminase activity in 3D organotypics. Taken together, these data suggests STS is important for regulating epidermal homeostasis and differentiation. The effect of STS knockdown on the transcriptome in primary keratinocytes was analyzed using RNA-Seq. Differentially expressed genes were grouped by pathogenic mechanism in relation to RXLI: differentiation, lipid metabolism, desquamation, testicular development, visual and behavioural disorders. Several genes were validated, including: ALDH1A1, ALDH3A1, ACER1, CYP4F22, OXTR, and UGCG. Novel data included the linking of ALDH1A1 and ALDH3A1 loss to corneal opacities, and OXTR loss to prolonged labour and behavioural disorders. Loss of ACER1 and UGCG will affect lipid metabolism in the epidermis. My hypothesis has been partially confirmed as many pathomechanisms observed in RXLI patient samples have been replicated in the 3D model, including many novel changes in protein expression. In conclusion, this thesis has identified novel pathomechanisms related to the clinical phenotype, both cutaneous and extracutaneous. This gives a greater understanding of the causative mechanisms of RXLI, and also provides a new explanation for the extracutaneous presenting features of RXLI.
AuthorsMcGeoghan, Farrell Thomas Sean
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