Fig. 2

Functional analysis of TRAPPC2 variants found in the SEDT-XL patients (a) Predicted amino acid sequences of the TRAPPC2 variants found in the SEDT-XL individuals. The c.1A > T missense variant would change the starting codon Methionine to Leucine. The deletion variant, c.40delG, would only produce the first 13 amino acids correctly and then abruptly terminate the translation. The gene accession number for human TRAPPC2 mRNA is indicated. b DNA constructs used in this study. Wild-type (WT) or the TRAPPC2 variants are expressed under the CMV promoter. HA tag is introduced at the C-terminus of the cDNAs and thus the HA tag does not affect translation of the TRAPPC2 variants. The arrows indicate primers for RT-PCR. These primers are specific to the vector used in this experiment to exclude any endogenous TRAPPC2 genes. c Chromatogram displaying the mutations in the TRAPPC2 expression vectors. A missense mutation at the position 1 showed a change of base from adenine to thymine in the left pair. A deletion of guanine at the position 40 is demonstrated in the right pair. d-e Analyses of the TRAPPC2 variant expression at protein and transcript level. 293 T HEK cells were transfected with individual pCDNA3.1, WT, or TRAPPC2 variant expressing vector, and cell lysates were subjected to Western blot analysis with HA antibody (d) and used to prepare total RNA for RT-PCR (e). The size markers for DNA fragments and proteins are indicated