As its name suggests, amyloid precursor protein (APP) gives rise to the infamous Aβ peptide that wreaks havoc in Alzheimer’s disease. Now, a study pinpoints a pathological role for non-amyloidogenic processing of APP. Researchers reported July 15 in Neuron that soluble APPα (sAPPα) runs amok in a mouse model of Fragile X Syndrome (FXS), a neurodevelopmental disorder. In young mice, the peptide ramps up the translation of other proteins, which triggers synaptic deficits and behavioral problems, according to the paper. The researchers, led by Claudia Bagni at KU Leuven in Belgium, traced the boost in sAPPα to a rise in the expression of APP and ADAM10, aka α-secretase, the protease that generates sAPPα from APP. They also found evidence that this pathway goes awry in human cases of FXS, and proposed that blocking ADAM10 in early life may help treat the disorder.
“This [study] is significant in that it raises the intriguing possibility that abnormal APP production and processing could underlie both Fragile X, a neurodevelopmental disorder, and Alzheimer’s disease, the most common neurodegenerative disease in old age,” commented Hui Zheng of Baylor College of Medicine in Houston. She added that a bevy of questions remain unanswered.
FXS is caused by the loss of or mutations in the gene encoding Fragile X mental retardation protein (FMRP), an RNA-binding protein that turns down the translation of many genes involved in neural function. About a third of people with FXS—who have behavioral, learning, and anxiety problems—also meet the criteria for Autism Spectrum Disorder (ASD) (see Lozano et al., 2014). APP is among the 40 or so genes confirmed to be regulated by FMRP (see Westmark and Malter, 2007; Pascuito and Bagni, 2014).
Soluble APPα promotes synaptogenesis and boosts synaptic density (see Bell et al., 2008; Tyan et al., 2012; Hick et al., 2015). Both APP and sAPPα are highly expressed during periods of intense synaptogenesis in humans and in mice (see Moya et al., 1994). People with FXS reportedly have an overabundance of immature dendritic spines, which are long, skinny, and twisted (see Irwin et al., 2011).
First author Emanuela Pascuito and colleagues wondered whether too much of a good thing, sAPPα, could explain the synaptic mayhem in FXS. The researchers tracked levels of APP, sAPPα, and ADAM10 throughout postnatal development in FMR1 knockout mice, which display many of the characteristics of the syndrome. APP levels were normal until the animals reached 3 weeks of age, at which point they shot up compared to controls and remained high throughout adulthood. While the total amount of APP rose in the KO neurons, cell surface levels of APP were lower than those in normal cells, suggesting internalization or an increased shedding of APP fragments. ADAM10 and sAPPα also increased in the knockout mice at 3 weeks, though their levels returned to normal by 3 months of age.
The researchers next zoomed in on dendritic spines. They compared the shape and size of the spines on neurons isolated from normal or FMR1 KO mice, and found that spines on knockout neurons tended to be more abundant and immature-looking; they were long and thin, rather than mushroom-shaped or stubby. Knocking down APP expression in these neurons reduced the density of these wispy spines, but they flourished again when the authors added sAPPα. This suggested that the elevated levels of sAPPα in FMR1-negative mice were responsible for the shift toward immature spines.
Because FMR1 KO mice are known to have elevated protein synthesis, the researchers next wanted to test whether elevated sAPPα helped promote this effect. They labeled newly synthesized proteins in cortical neurons from KO versus normal mice and found that, as expected, the former pumped out more protein. However, when the researchers crossed the knockout mice with animals expressing only a single copy of APP or ADAM10, cortical neurons from the offspring made normal amounts of protein. On the other hand, adding sAPPα to neurons from any of the mouse strains boosted protein synthesis. Together, these results suggested that sAPPα helped drive exaggerated protein synthesis in the Fragile X mice.
How would elevated sAPPα raise global protein synthesis? The mGluR5 pathway is a possibility. Previous reports indicate that this metabotropic glutamate receptor and downstream MAP kinase signaling are affected in FXS, and that enhanced signaling through this receptor elevates protein synthesis. The researchers found that sAPPα enhanced MAP kinase phosphorylation of ERK1/2 in cortical neurons, and that this increase was abolished when the cells were treated with a GluR1/5 inhibitor. Bagni said that her lab is currently investigating how sAPPα switches on the GluR5 pathway.
Would inhibiting ADAM10, and thus production of sAPPα, ameliorate FXS symptoms in the mice? To find out, the researchers injected the animals with Tat-Pro709-729 ADAM10, a peptide fragment that contains part of the ADAM10 intracellular domain and interferes with its protease activity. Tat-Pro lowered sAPPα levels to normal in FMR1 KO mice. The inhibitor also normalized long-term depression (LTD), which is enhanced in FXS. Tat-Pro also reversed several learning and behavioral problems in the knockouts, including working memory deficits, hyperactivity, and poor nest building.
sAPPαOverdrive. In wild-type neurons (left), FMRP keeps protein translation in check. In FXS neurons, loss of FMRP takes the brakes off translation, while overproduction of sAPPα keeps the translational machinery humming. [Courtesy of Pascuito et al., Neuron 2015.]
“This is a great paper,” commented Bernadette Allinquant at INSERM in Paris. Allinquant has worked to characterize the functions of sAPPα for decades. “I’ve always thought of sAPPα as a ‘good molecule.’ It has neurotrophic and neuroprotective properties and increases LTP,” she said. “For the first time, this paper reveals that too much sAPPα at the wrong time is a bad thing.”
The researchers searched for clues that this pathway played a role in FXS in humans. They found elevated APP (but not ADAM10) in brain and lymphoblastoid cells from older patients, and elevated levels of both APP and ADAM10 in fibroblasts from younger patients. These changes paralleled the developmental regulation of these proteins in FMR1 KO mice—with APP levels high throughout adulthood, and ADAM10 only going up during younger life.
The researchers are collecting fibroblast samples, and plan to measure fluctuations in sAPPα and ADAM10 from early childhood into adulthood. Bagni wants to home in on the window during which sAPPα rises in people with FXS, and test whether giving an ADAM10 inhibitor during that time could ameliorate some symptoms of the disease. “After that critical period, ADAM10 is no longer dysregulated, and this treatment would not work,” she said. She also plansto test whether treatment of juvenile mice with the inhibitor fends off the manifestation of disease later on.
Recent studies reported elevated serum sAPPα in autism spectrum disorder and FXS patients, and Bagni envisions using such a biomarker to select and monitor patients in clinical trials (see Ray et al., 2011; Erickson et al., 2014). The elevated sAPPα in autism patients who do not have FXS raises the interesting possibility that the fragment plays a role in a host of disorders.
“These studies contribute to a growing body of knowledge suggesting that APP processing is developmentally regulated and aberrant in both fragile X and autism,” wrote Cara Westmark of the University of Wisconsin in Madison. She added that much is left to learn about the changing balance between non-amyloidogenic and amyloidogenic APP processing throughout early development and adulthood. She also cautioned that toying with ADAM10 activity or levels of APP metabolites could promote seizures, as has been demonstrated in ADAM10 knockout mice. Bagni said that the ADAM10 inhibitors merely return the protease activity to normal.—Jessica Shugart
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