As the γ-secretase complex has given up some of its secrets, it has created new puzzles to solve, such as how many variations of the complex exist in vivo, and how these variations might impact γ-secretase function from tissue to tissue and from substrate to substrate (see ARF related news story). One variable is Aph1 (see ARF related news story), which appears to serve as a founding member of the complex by binding with nicastrin. Two recent knockout studies reveal that the Aph1A gene is the only one of the three versions in mice that is critical for proper ontogenetic development, solidifying the case for distinct γ-secretase complexes.

The new insights about Aph1 come from a study reported in the January 5 issue of The Journal of Neuroscience by Philip Wong, first author Guojoun Ma, and colleagues at Johns Hopkins University in Baltimore, Maryland, along with a new study published online Jan 21 in PNAS by Bart De Strooper, first author Lutgarde Serneels, and colleagues at the Flanders Interuniversity for Biotechnology in Leuven, Belgium, and Radboud University in Nijmegen, the Netherlands. The latter group generated knockouts of Aph1B and Aph1C, as well as BC double knockouts, in addition to the Aph1A KOs.

While knocking out one copy of the Aph1A gene caused no gross abnormalities, both studies found that the Aph1A -/- mouse was a real disaster. No animals “survived” to embryonic day 11. However, the phenotype was not as drastic as that seen when there is a full loss of γ-secretase activity (i.e., PS1- plus PS2-deficient mice, or nicastrin-deficient mice) or mice lacking the gene for the developmental regulator Notch1, a γ-secretase substrate. Although there were major disruptions in angiogenesis, the Aph1A-deficient mouse embryos were able to develop caudal morphologic features such as the beginnings of hind limbs and a tail, indicating that Aph1B and C were able to compensate to some degree for loss of the A isoform. Still, Aph1A emerges as the major developmental isoform, as Serneels and colleagues report that Aph1B-deficient, Aph1C-deficient, and Aph1B/C-deficient mice all grow to adulthood and reproduce apparently normally.

Using embryonic fibroblasts from these knockout mice, Serneels looked into how the Aph1 deficiencies affected γ-secretase cleavage. As indicated by assays of endogenous APP and N-cadherin—γ-secretase substrates that will pile up in the absence of γ-secretase activity—only the Aph1A knockout noticeably impacts γ-secretase cleavage in the embryonic fibroblasts. Interestingly, there seems to be no favoritism played between APP vs. Notch processing. Serneels and colleagues found that the lack of Aph1A inhibited both APP and Notch γ-secretase cleavage by about 70 percent.

An echo of the major disruption of γ-secretase activity was found by Ma and colleagues in their own Aph1A-/- fibroblasts. They noted significant reductions in the formation of high-molecular-weight γ-secretase complexes, along with reductions in mature nicastrin, presenilin, and PEN-2. Ma and colleagues note that the reduction in mature nicastrin levels in Aph1A-deficient fibroblasts would appear to support the notion that Aph1 is necessary to stabilize nicastrin, forming an initial "pre-complex" on the road to γ-secretase (see Gu et al., 2003; Takasugi et al., 2003; Hu and Fortini, 2003; LaVoie et al., 2003).

The end result of these disruptions, both research groups found, was a significant reduction in Aβ accumulation in Aph1A-deficient fibroblasts transfected with human APP. However, when Ma and colleagues expressed the different isoforms of Aph1—Aph1A in its long (L) and short (S) splice variants, as well as Aph1B, and Aph1C—in Aph1A-/- fibroblasts, they found that all were able to significantly increase levels of secreted Aβ40. "Our demonstration that all murine APH-1 isoforms can complement the Aph-1a null phenotype in fibroblasts supports the idea that the four murine APH-1 isoforms (APH-1aL, APH-1aS, APH-1b, and APH-1c) coupled with PS1 and PS2 define a set of eight distinct functional γ-secretase complexes," write Ma and colleagues. In this light, it is of interest that Serneels and colleagues found differences in γ-secretase-mediated APP processing (as inferred from levels of APP-CTF, as well as PS, nicastrin, and PEN2) in different brain regions in mice lacking both Aph1B and C. For example, there was a twofold accumulation of APP-CTF in brainstem and olfactory bulb. This would suggest that Aph1B in humans, presumed to be the equivalent of the closely related murine B and C variants, is more active in some regions than others, supporting the notion that different Aph1 isoforms give rise to different γ-secretase complexes.

"The identification of a specific APH-1 isoform selectively expressed in the CNS during aging would have important therapeutic implications for efforts to selectively alter γ-secretase activity to ameliorate β-amyloidosis in AD," write Ma and colleagues. For their part, Serneels and colleagues consider the possibility that the survival of Aph1BC-deficient mice makes BC (or B in humans) a safer, and thus prime drug target for AD, but they also point to behavioral and physiological abnormalities in a rat model of reduced Aph1BC expression that caution against undue optimism in this approach.—Hakon Heimer


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News Citations

  1. St. Moritz, Part 4. (It) Take(s) Five (To Make Aβ), New BACE Mice
  2. Homing in on Roles for PS Complex Proteins APH-1 and PEN-2

Paper Citations

  1. . APH-1 interacts with mature and immature forms of presenilins and nicastrin and may play a role in maturation of presenilin.nicastrin complexes. J Biol Chem. 2003 Feb 28;278(9):7374-80. PubMed.
  2. . The role of presenilin cofactors in the gamma-secretase complex. Nature. 2003 Mar 27;422(6930):438-41. PubMed.
  3. . Different cofactor activities in gamma-secretase assembly: evidence for a nicastrin-Aph-1 subcomplex. J Cell Biol. 2003 May 26;161(4):685-90. PubMed.
  4. . Assembly of the gamma-secretase complex involves early formation of an intermediate subcomplex of Aph-1 and nicastrin. J Biol Chem. 2003 Sep 26;278(39):37213-22. PubMed.

Further Reading

Primary Papers

  1. . APH-1a is the principal mammalian APH-1 isoform present in gamma-secretase complexes during embryonic development. J Neurosci. 2005 Jan 5;25(1):192-8. PubMed.
  2. . Differential contribution of the three Aph1 genes to gamma-secretase activity in vivo. Proc Natl Acad Sci U S A. 2005 Feb 1;102(5):1719-24. PubMed.