Novel genomic approach reveals gene mutation isn’t simple answer.
Researchers at the University of California, San Diego School of Medicine have used precise genetic engineering of human induced pluripotent stem cells to dissect how a key mutated protein contributes to familial Alzheimer’s disease (AD). Their results show that a straightforward loss-of-function of the protein does not explain the inherited form of the disease, a finding that refines our understanding of familial AD biology and will influence future drug development efforts.
The study, published in Cell Reports, was led by Lawrence Goldstein, PhD, professor in the Departments of Cellular and Molecular Medicine and Neurosciences and director of the UC San Diego Stem Cell Program. Goldstein and colleagues combined stem cell biology with genomic editing to create isogenic human neurons that differ only at the presenilin 1 (PS1) gene, enabling them to assign precise functional consequences to specific mutations.
Familial Alzheimer’s disease represents a smaller, inherited portion of early-onset AD cases, driven by mutations in genes such as presenilin 1 (PS1) and others. The majority of Alzheimer’s cases are sporadic, influenced by age and other risk factors rather than a single inherited mutation. Understanding how familial mutations alter cellular processes can illuminate core mechanisms that may also operate, in different ways, in sporadic disease.
PS1 is an essential component of the gamma-secretase complex, a membrane-associated protease that cleaves many type‑1 transmembrane proteins. One of the best-known substrates of gamma-secretase is amyloid precursor protein (APP). When gamma-secretase cleaves APP it produces short peptide fragments called amyloid beta. Certain forms of amyloid beta can aggregate and form the plaques historically associated with AD pathology. However, the relationship between PS1 mutations, gamma-secretase activity, amyloid beta generation, and neurodegeneration is complex and has remained controversial.
Using human neurons derived from induced pluripotent stem cells (iPSCs), the UC San Diego team edited the PS1 gene to create cells that carried specific familial AD mutations in an otherwise identical genetic background. The source iPSCs included well-characterized lines originating from the genome of noted biologist Craig Venter. Working with isogenic cell lines allowed the investigators to isolate the effects of single PS1 mutations without confounding background genetic differences.
The experiments revealed that PS1 mutations do not produce a simple total loss of protein function. Instead, the mutations altered the “molecular scissors” activity of gamma-secretase in a more nuanced way: they increased the frequency of aberrant cleavage events that produce potentially harmful amyloid beta fragments. Goldstein and colleagues observed that these mutations roughly doubled the incidence of such undesirable cleavages compared with wild-type PS1 in their system. In normal cells, gamma-secretase performs most cleavages without producing harmful fragments; the familial mutations increase the proportion of cuts that yield problematic amyloid beta species.
To ensure that the results reflected the intended edits and not off-target effects of genome engineering, the investigators performed whole exome sequencing on the engineered cell lines. Co-author Kun Zhang, PhD, associate professor in the Department of Bioengineering at UC San Diego, reported that the sequencing comparisons confirmed the genome editing approach did not introduce additional mutations that could confound the interpretation.
These findings narrow the range of possible mechanisms by which PS1 mutations promote familial AD. By demonstrating altered gamma-secretase processing rather than total loss of PS1 function, the study helps refine therapeutic strategies: instead of attempting to restore a non-existent global activity, effective interventions may need to correct or prevent the specific, aberrant cleavage events that generate toxic amyloid beta fragments.
Notes about this Alzheimer’s disease and neurogenetics research
Lead investigators and co-authors include Lawrence S.B. Goldstein, Kun Zhang, Grace Woodruff, Jessica E. Young, Fernando J. Martinez, Floyd Buen, Jennifer Kinaga, Athurva Gore, Zhe Li, and Shauna H. Yuan, representing the Departments of Cellular and Molecular Medicine, Neurosciences, and Bioengineering, along with the Institute for Genomic Medicine and the Institute of Engineering in Medicine at UC San Diego.
Funding support for this research came in part from the California Institute for Regenerative Medicine, the National Institutes of Health and the National Institute on Aging (grant R01AG032180), and the A.P. Giannini Foundation for Medical Research.
Contact: Scott LaFee – UCSD
Source: UCSD press release; original research published in Cell Reports (2013) titled “The Presenilin-1 ΔE9 Mutation Results in Reduced γ-Secretase Activity, but Not Total Loss of PS1 Function, in Isogenic Human Stem Cells” by Grace Woodruff et al., doi:10.1016/j.celrep.2013.10.018
Keywords: familial Alzheimer’s disease, presenilin 1, PS1, gamma-secretase, amyloid beta, induced pluripotent stem cells, isogenic human neurons, UC San Diego, neurogenetics, Alzheimer’s research.