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. 2023 Dec 11;12:RP89891. doi: 10.7554/eLife.89891

Figure 2. UFMylation pathway, conservation of UBA5, and generation of fly Uba5 loss of function (LoF) alleles.

Figure 2.

(A) Alignment of the human UBA5 and fly Uba5 protein sequences. The functional domains of UBA5 are marked in colored boxes. The developmental and epileptic encephalopathy 44 (DEE44)-associated variants are marked in the protein topology diagram and the protein sequence alignment (letters in red). (B) Generation of the Uba5T2A-Gal4 allele and the uses of the allele in flippase (FLP)-mediated conversion. The expression of the GAL4 to drive a fluorescent protein allows assessment of gene expression, and humanization of the flies by expression of human UBA5 cDNA. (C) Generation of Uba5 null allele by CRISPR-mediated indel formation. (D) Loss of Uba5 causes lethality in early developmental stage. The lethality is rescued by a genomic rescue construct, the expression of FLP (Uba5T2A-Gal4 mutants only), and the expression of human UBA5 cDNA.