In the HEK293T human embryonic kidney cells, none of the constructs led to sufficient levels of expression above the control plasmid. These results were not unexpected, because studies of β3 subunit expression have been challenging due to its low level of endogenous expression in a limited number of certain cell types (Cauley et al., 1996). Fortunately, we were able to compare expression levels of the five constructs with control plasmid lacking any inserts using two cell lines: P19 mouse embryonal carcinoma cells and SH-SY5Y human neuroblastoma cells. In the P19 cells, there was a significant difference in expression between the long construct containing the common haplotype and the minor haplotype (Figure 3a) providing evidence that allelic variation in this promoter region may be associated with gene expression. There were no significant differences in expression among any of the constructs in the undifferentiated SH-SY5Y cells (Figure 3b). However, there was a significant difference between the long common and mixed haplotypes, and suggestive evidence for a difference between the long common and minor haplotypes. This last comparison probably failed to reach significance because of the variability in signal that is obtained from these assays.
