A paper published January 29 in Cell Reports claims that among the earliest changes in sporadic Alzheimer’s disease (SAD) are genetic changes that lead to premature neurogenesis. Scientists led by Bruce Yankner, Harvard Medical School, found that neural progenitor cells derived from fibroblasts of patients with late-onset AD have nuclei bereft of the repressor element 1-silencing transcription factor. REST normally reins in neuronal differentiation. The result? Unchecked differentiation and depletion of the neural progenitor pool.
- Neural progenitors from AD patients lack the nuclear repressor REST.
- Without it, neural differentiation accelerates, prematurely depleting the stem cell pool.
- ApoE4 alone mimics this phenotype.
What’s more, the same phenotype arises in progenitor cells engineered to express the AD risk allele ApoE4. The authors saw abnormalities in the nuclear membranes of these differentiated cells, implying that its disruption is also part of early AD. Together, the results point to a new set of cellular changes in sporadic AD pathogenesis.
“This suggests that the changes to the genome may be happening at an early time point,” said Bess Frost, University of Texas Health, San Antonio, who was not involved in the study. “I’m excited because it provides even more evidence that there’s this problem with overall genomic architecture in the context of Alzheimer’s disease.”
Yankner had previously implicated REST in aging, when his lab reported that REST is highly expressed in aging and correlates with better cognitive function even in the face of AD pathology (Mar 2014 news). The transcriptional repressor turns down expression of genes for cell death and Alzheimer’s pathology, and turns up genes that protect against oxidative damage. Conversely, postmortem neuronal nuclei from AD dementia patients lacked REST, and the protein was down by half in people who had died with mild cognitive impairment.
“Now in sporadic Alzheimer’s disease neurons [derived from iPSCs, Yankner] was able to recapitulate this, which really validates his previous observations,” said Li-Huei Tsai, Massachusetts Institute of Technology, Cambridge. Tsai is a co-author on the paper, though she contributed mostly technical support. “The pathways underlying this phenotype in this cell culture system may be very fundamental to the disease mechanism and REST could be contributing, though I wouldn’t be surprised if there were other factors involved in this process of accelerated neurodifferentiation and reduced progenitor cell renewal,” she said.
Co-first authors Katharina Meyer and Heather Feldman went looking for fundamental cellular changes in SAD by examining induced pluripotent stem cells (iPSCs) from patients. Past studies on iPSCs from AD patients primarily focused on mutations that cause early onset disease (Muratore et al., 2014; Sproul et al., 2014; Yagi et al., 2011). While some studies have included iPSCs from sporadic patients, those haven’t settled on a common phenotype or central mechanism (Israel et al., 2012; Kondo et al., 2013).
For this study, researchers collected dermal fibroblasts of five people with sporadic AD and six age-matched controls, reverted the cells to a pluripotent state, and then derived neural progenitors. This type of cell gives rise to most of the differentiated cells of the human brain, including astrocytes, oligodendrocytes, and mature neurons.
Right away, the researchers saw differences between the transcription profiles of neural progenitors from SAD and controls. The former expressed more genes related to neurogenesis and neuronal differentiation. Their tau expression was upregulated, and ApoE expression was down. A greater proportion of SAD cells differentiated into neurons; fewer proliferated to keep the pool of neural progenitors going.
Were there differences among mature neurons? To find out, the authors differentiated the neuronal progenitors into neurons for six weeks. During that time, SAD cells matured faster, some in less than half the time of control cells. Action potentials appeared in the SAD cells weeks earlier than in control cells, occurring twice as often and reaching higher amplitudes, suggesting they were hyperexcitable. SAD cells expressed more genes for synapse formation, axon guidance, neurotransmitter receptors, and ion channels.
What drives these expression differences? A search of the ENCODE chromatin immunoprecipitation (ChIP) database revealed that this particular suite of genes is normally governed by the transcriptional repressor REST, a kind of brake on their expression. Without REST, these genes go into hyper-expression mode. The gene analysis also turned up two components of the polycomb repressive complex 2 (PRC2), EZH2 and SUZ12, which interact with REST to help suppress expression.
REST bound less to its target genes in SAD cells. While these contained as much REST mRNA and protein as control cells, the normally nuclear protein seemed stuck in the cytoplasm. However, when Meyer overexpressed REST in SAD progenitor cells, they differentiated less and expressed normal levels of REST target genes.
Given that ApoE4 is the biggest genetic risk factor for AD, the authors wondered whether its expression would bring on this phenotype in iPSCs derived from people without AD. The authors used CRISPR-Cas9 to convert a homozygous ApoE3 iPSC control line to a homozygous ApoE4 one. This drove up expression of neural differentiation genes and of tau in neural progenitors. As in the SAD cells, REST strayed from the nucleus into the cytoplasm. Conversely, editing an ApoE4 iPSC line to express only ApoE3 led to lower expression of those genes. Together, the results suggest that neural progenitors in people with SAD are pushed to differentiate early, and ApoE4 alone can bring on this phenotype.
“In adults, if you have accelerated differentiation of neural progenitors, you’ll eventually exhaust the stem cell pool,” Yankner explained. “By the time someone is aging, we would hypothesize that their ability to generative new neurons would be diminished.” A previous study suggested that accelerated differentiation disrupts memory in mice (Akers et al., 2014). “It could be that these neurons don’t function as well,” said Yankner.
If REST is wandering out into the cytoplasm, could there be a problem with nucleocytoplasmic transport? Under a confocal microscope, the authors observed that SAD cells took on an unusual shape, with folds and circular structures within the nuclear lamina (see image below). These abnormal indentations resemble ones recently linked to tau pathology, for example in iPSC-derived neurons from frontotemporal dementia patients, and in AD postmortem brain (Jan 2019 news; Frost et al., 2016; Sep 2018 news). Scientists also reported abnormal nucleocytoplasmic transport in transgenic mouse models of AD. No one has previously seen the circular structures Yankner observed in the current study. He plans to do three-dimensional imaging of these nuclei to visualize them more fully.
“There is this converging set of observations in the area of neurodegenerative diseases that there’s a fundamental problem with the nuclear membrane/lamina, and the regulation of nucleocytoplasmic transport, which may contribute to the neurodegenerative process,” Yankner told Alzforum.
“These are interesting results,” said Robert Rissman, University of California, San Diego. “REST is an important protein and regulator of many different genes. I think this is a potential mechanism in AD neuropathology.” Rissman previously reported reduced REST in neuronal exosomes found in the blood of people progressing from mild cognitive impairment to AD dementia (Winston et al., 2016). It would be worth checking if REST levels vary in exosomes released from the iPSCs in Yankner’s study, given that they reflect an earlier stage of disease, he added.—Gwyneth Dickey Zakaib