The Notch cleavage efficiency of Aph aS containing complexes
The Notch-cleavage efficiency of Aph1aS containing complexes was lower than that of Aph1aL γ-secretases, especially in combination with PS2, suggesting that Aph1aS is a determinant of low Notch-cleavage specificity, considering that the different subcellular localization of PS1 and PS2 didn't affect endogenous NICD production . Saito et al. reported that mRNA level of Aph1aS was 1.5∼3-fold higher than that of Aph1aL in HEK293 (∼3-fold), SH-SY5Y (∼2.5-fold), Hela (1.5∼2-fold) and human adult RN486 (2.5∼3-fold) , though it is unknown which combinations of γ-secretase are formed most prominently. It was reported that Aph1b γ-secretases might be a potential target of γ-secretase inhibitors for AD treatment because of their small side effects in knock-out models , whereas Aph1a γ-secretases shouldn't be the target because of their important role in development . Our results, however, revealed that Aph1aS γ-secretase cleaved Notch less efficiently than Aph1aL, suggesting that it is rather Aph1aS that should be targeted. In contrast, according to our data specific inhibition of Aph1b γ-secretases might cause side effect because of equal inhibition of APP and Notch signalling. Previous different conclusion that Aph1b γ-secretases play less important roles in Notch-cleavage compared to Aph1a , might be explained as follows. Knockdown of Aph1b is reported not to cause reduction of the expression levels of other γ-secretase components, implying that Aph1a γ-secretases still can work properly. In contrast, knockdown of Aph1a caused marked reduction of expression level of the other γ-secretase components , , suggesting that Aph1b γ-secretase formation is also affected.
In this manuscript, we showed that cells expressing PS2/Aph1aS γ-secretase produced Aβ with higher Aβ42(43)/40 ratio and lower Notch-cleavage efficiency. However, this cell line contains only a slight amount of PS1/Aph1aS γ-secretase because cells expressing PS2 γ-secretase showed residual expression of PS1 and vice versa, but importantly almost no residual Aph1 expression (Supplementary Fig. S1). PS1/Aph1aS γ-secretase showed lower Aβ42(43)/40 ratio and higher Notch-cleavage efficiency than PS2/Aph1aS γ-secretase. Therefore, pure PS2/Aph1aS γ-secretase would be expected to show even higher Aβ42(43)/40 ratio and even lower Notch.
In conclusion, we here show that the six distinct human γ-secretases have different cleavage-efficiencies for Notch and differ in the Aβ species they produce, though it is needed to analyse Notch-cleavage and Aβ production by the distinct γ-secretases in neuronal cells to confirm that our findings are the case with the brain. Recent studies revealed the PS1/Aph1aL γ-secretase structure . Further analysis of different γ-secretase structure might aid in the development of structure-based specific γ-secretase inhibitors. Targeting the PS2/Aph1aS γ-secretase complex would be expected to have the greatest effect of reducing Aβ42(43)/Aβ40 with the fewest side effects caused by inhibition of Notch signalling.
Competing financial interests
Y.Y. was supported by Japan Society for the Promotion of Science. This work was supported in part by an Intramural Research Grant (MHLW; 26-8) for Neurological and Psychiatric Disorders of NCNP, a research grant for Comprehensive Research on Disability Health and Welfare from the Ministry of Health, Labour and Welfare (MHLW; H26-Sinkeikinn-ippan-004) and a Grant-in-Aid from the MHLW of Japan (to S.I.).
Alzheimer’s disease (AD) is a devastating neurological disorder and is currently estimated to affect 5.3million Americans. In 2015, AD and other related dementias will cost the United States $226billion, and if the current disease trajectory is maintained, the associated costs could rise to as high as $1.1trillion by 2050. AD is the only top ten cause of death in America that cannot be prevented, cured or slowed, and thus there is an urgent medical need for disease modifying therapeutic agents.