Tight control of the APP-Mint1 interaction in regulating amyloid production.

Generation of amyloid-ß (Aß) peptides through the proteolytic processing of the amyloid precursor protein (APP) is a pathogenic event in Alzheimer's disease (AD). APP is a transmembrane protein and endocytosis of APP mediated by the YENPTY motif is a key step in Aß generation. Mints, a family of cytosolic adaptor proteins, directly bind to the YENPTY motif of APP and facilitate APP trafficking and processing. Here, we generated and examined two Mint1 mutants, Tyr633Ala of Mint1 (Mint1Y633A) that enhanced APP binding, and Tyr549Ala and Phe610Ala mutant (Mint1Y549A/F610A), that reduced APP binding. We investigated how perturbing the APP-Mint1 interaction through these Mint1 mutants alter APP and Mint1 cellular dynamics and Mint1's interaction with its other binding partners. We found that Mint1Y633A increased binding affinity specifically for APP and presenilin1 (catalytic subunit of γ-secretase), that subsequently enhanced APP endocytosis in primary murine neurons. Conversely, Mint1Y549A/F610A exhibited reduced APP affinity and Aß secretion. The effect of Mint1Y549A/F610A on Aß release was greater compared to knocking down all three Mint proteins supporting the APP-Mint1 interaction is a critical factor in Aß production. Altogether, this study highlights the potential of targeting the APP-Mint1 interaction as a therapeutic strategy for AD.

Bisphenol A promotes stress granule assembly and modulates the integrated stress response.

Bisphenol-A (BPA) is a ubiquitous precursor of polycarbonate plastics that is found in the blood and serum of >92% of Americans. While BPA has been well documented to act as a weak estrogen receptor (ER) agonist, its effects on cellular stress are unclear. Here, we demonstrate that high-dose BPA causes stress granules (SGs) in human cells. A common estrogen derivative, ß-estradiol, does not trigger SGs, indicating the mechanism of SG induction is not via the ER pathway. We also tested other structurally related environmental contaminants including the common BPA substitutes BPS and BPF, the industrial chemical 4-nonylphenol (4-NP) and structurally related compounds 4-EP and 4-VP, as well as the pesticide 2,4-dichlorophenoxyacetic acid (2,4-D). The variable results from these related compounds suggest that structural homology is not a reliable predictor of the capacity of a compound to cause SGs. Also, we demonstrate that BPA acts primarily through the PERK pathway to generate canonical SGs. Finally, we show that chronic exposure to a low physiologically relevant dose of BPA suppresses SG assembly upon subsequent acute stress. Interestingly, this SG inhibition does not affect phosphorylation of eIF2α or translation inhibition, thus uncoupling the physical assembly of SGs from translational control. Our work identifies additional effects of BPA beyond endocrine disruption that may have consequences for human health.

Targeting the APP-Mint2 Protein-Protein Interaction with a Peptide-Based Inhibitor Reduces Amyloid-ß Formation.

There is an urgent need for novel therapeutic approaches to treat Alzheimer's disease (AD) with the ability to both alleviate the clinical symptoms and halt the progression of the disease. AD is characterized by the accumulation of amyloid-ß (Aß) peptides which are generated through the sequential proteolytic cleavage of the amyloid precursor protein (APP). Previous studies reported that Mint2, a neuronal adaptor protein binding both APP and the γ-secretase complex, affects APP processing and formation of pathogenic Aß. However, there have been contradicting results concerning whether Mint2 has a facilitative or suppressive effect on Aß generation. Herein, we deciphered the APP-Mint2 protein-protein interaction (PPI) via extensive probing of both backbone H-bond and side-chain interactions. We also developed a proteolytically stable, high-affinity peptide targeting the APP-Mint2 interaction. We found that both an APP binding-deficient Mint2 variant and a cell-permeable PPI inhibitor significantly reduced Aß42 levels in a neuronal in vitro model of AD. Together, these findings demonstrate a facilitative role of Mint2 in Aß formation, and the combination of genetic and pharmacological approaches suggests that targeting Mint2 is a promising therapeutic strategy to reduce pathogenic Aß levels.

A rare autism-associated MINT2/APBA2 mutation disrupts neurexin trafficking and synaptic function.

MINT2/APBA2 is a synaptic adaptor protein involved in excitatory synaptic transmission. Several nonsynonymous coding variants in MINT2 have been identified in autism spectrum disorders (ASDs); however, these rare variants have not been examined functionally and the pathogenic mechanisms are unknown. Here, we examined the synaptic effects of rat Mint2 N723S mutation (equivalent to autism-linked human MINT2 N722S mutation) which targets a conserved asparagine residue in the second PDZ domain of Mint2 that binds to neurexin-1α (Nrxn1α), a presynaptic cell-adhesion protein implicated in ASDs. We show the N723S mutation impairs Nrxn1α stabilization and trafficking to the membrane while binding to Nrxn1α remains unaffected. Using time-lapse imaging in primary mouse neurons, we found that the N723S mutant had more immobile puncta at neuronal processes compared to Mint2 wild type. We therefore, reasoned that the N723S mutant may alter the co-transport of Nrxn1α at axonal processes to presynaptic terminals. Indeed, we found the N723S mutation affected Nrxn1α localization at presynaptic terminals which correlated with a decrease in Nrxn-mediated synaptogenesis and miniature event frequency in excitatory synapses. Together, our data reveal Mint2 N723S leads to neuronal dysfunction, in part due to alterations in Nrxn1α surface trafficking and synaptic function of Mint2.

CLASP2 Links Reelin to the Cytoskeleton during Neocortical Development.

The Reelin signaling pathway plays a crucial role in regulating neocortical development. However, little is known about how Reelin controls the cytoskeleton during neuronal migration. Here, we identify CLASP2 as a key cytoskeletal effector in the Reelin signaling pathway. We demonstrate that CLASP2 has distinct roles during neocortical development regulating neuron production and controlling neuron migration, polarity, and morphogenesis. We found downregulation of CLASP2 in migrating neurons leads to mislocalized cells in deeper cortical layers, abnormal positioning of the centrosome-Golgi complex, and aberrant length/orientation of the leading process. We discovered that Reelin regulates several phosphorylation sites within the positively charged serine/arginine-rich region that constitute consensus GSK3ß phosphorylation motifs of CLASP2. Furthermore, phosphorylation of CLASP2 regulates its interaction with the Reelin adaptor Dab1 and this association is required for CLASP2 effects on neurite extension and motility. Together, our data reveal that CLASP2 is an essential Reelin effector orchestrating cytoskeleton dynamics during brain development.

Discerning differences: gastroesophageal reflux and gastroesophageal reflux disease in infants.

Gastroesophageal reflux (GER) is a frequently encountered problem in infancy; it commonly resolves spontaneously by 12 months of age. Caregivers are challenged to discriminate between physiologic GER and the much less common and more serious condition of pathologic gastroesophageal reflux disease (GERD). Pathologic GERD may require more extensive clinical evaluation and necessitate treatment. GERD may be primary or secondary; secondary GERD is associated with a number of genetic syndromes, chromosomal abnormalities, birth defects, or a host of neurologic conditions frequently seen in the newborn intensive care unit. This article reviews the unique anatomic, physiologic, developmental, and nutritional vulnerabilities of infants that make them susceptible to GER and GERD. The North American Society of Pediatric Gastroenterology and Nutrition have recently developed a comprehensive evidence-based clinical practice guideline that structures the diagnostic approach and treatment option in infants with suspected and confirmed GERD. These guidelines provide clear definitions of GER and GERD to aid the clinician in distinguishing between the 2 conditions. They emphasize the use of history and physical examination and discuss the indications for the use of other diagnostic procedures, such as upper gastrointestinal studies, nuclear medicine scintiscan, esophagogastroduodenoscopy with biopsy, and esophageal pH probe monitoring. Management of GERD begins with a nonpharmacologic approach; the emphasis is on positioning, a trial of a hypoallergenic formula, and thickening of feedings. When these measures fail to control symptoms, a trial of either histamine(2) antagonists or a proton pump inhibitor may be indicated. Finally, surgical treatment may be needed if all other management measures fail. New sleep recommendations for infants with GERD are now consistent with the American Academy of Pediatrics' standard recommendations. Prone sleep positioning is only considered in unusual cases, where the risk of death and complications from GERD outweighs the potential increased risk of sudden infant death syndrome (SIDS). The nursing care of infants with GER and GERD, as well as relevant issues for parent education and support, are reviewed and are essential elements in managing this common condition.