Angiotensin-converting enzyme 2 (ACE2) is the main entry point in airway epithelial cells for SARS-CoV-2. ACE2 binding to the SARS-CoV-2 protein spike triggers viral fusion with the cell plasma ...membrane, resulting in viral RNA genome delivery into the host. Despite ACE2's critical role in SARS-CoV-2 infection, full understanding of ACE2 expression, including in response to viral infection, remains unclear. ACE2 was thought to encode five transcripts and one protein of 805 amino acids. In the present study, we identify a novel short isoform of ACE2 expressed in the airway epithelium, the main site of SARS-CoV-2 infection. Short ACE2 is substantially upregulated in response to interferon stimulation and rhinovirus infection, but not SARS-CoV-2 infection. This short isoform lacks SARS-CoV-2 spike high-affinity binding sites and, altogether, our data are consistent with a model where short ACE2 is unlikely to directly contribute to host susceptibility to SARS-CoV-2 infection.
Phosphoinositides are membrane phospholipids involved in a variety of cellular processes like growth, development, metabolism, and transport. This review focuses on the maintenance of cellular ...homeostasis of phosphatidylinositol 4,5-bisphosphate (PIP
2
), and phosphatidylinositol 3,4,5-trisphosphate (PIP
3
). The critical balance of these PIPs is crucial for regulation of neuronal form and function. The activity of PIP
2
and PIP
3
can be regulated through kinases, phosphatases, phospholipases and cholesterol microdomains. PIP
2
and PIP
3
carry out their functions either indirectly through their effectors activating integral signaling pathways, or through direct regulation of membrane channels, transporters, and cytoskeletal proteins. Any perturbations to the balance between PIP
2
and PIP
3
signaling result in neurodevelopmental and neurodegenerative disorders. This review will discuss the upstream modulators and downstream effectors of the PIP
2
and PIP
3
signaling, in the context of neuronal health and disease.
Type I interferons were not present in sufficient quantity to measure; however, the finding of lower CXCL-10 quantities in asthmatic explants was consistent with deficient innate antiviral defences ...in asthma. Because subjects with asthma in our study were on regular ICSs, the effects on influenza susceptibility of which are unknown, we also sought to determine whether the differences seen in the primary comparator groups reflected the effects of FP or disease. Parameter Healthy controls Subjects with moderate asthma P value N 10 10 - Age (y) 27 (22.5-48.0) 35.5 (23.3-48.5) .647∗ Sex (M/F) 6/4 6/4 1.0† FEV1 4.23 (3.03-5.14) 3.55 (2.12-4.59) .142∗ FEV1 % predicted 109.1 (99.6-112.2) 95.9 (63.1-108.0) .017∗ Inhaled steroid dose (μg/d) NA 280 (165.4-394.6) <.001∗ SABA (μg/d) NA 140 (89.98-190.00) <.001∗ Blood eosinophils (%) 2.87 (1.69-5.87) 3.15 (2.04-4.89) .986∗ Total blood IgE 33.3 (7.1-63.5) 89.6 (31.0-179.5) .023∗ Atopy (yes) 3/10 8/10 .025† Feno 18.25 (11.38-33.38) 33.75 (21.50-100.10) .027∗ Sputum eosinophils (%) 0 2.39 (0-3.68) .045∗ Sputum neutrophils (%) 11.4 (7.1-15.7) 64.5 (21.9-74.2) .006∗ Table E2List of antibody clones and fluorophores used for detection of cell types and activation markers on cells dispersed from bronchial biopsy explantsAPC, Antigen-presenting cell; EpCAM, epithelial cell adhesion molecule; FITC, fluorescein isothiocyanate; NA, not available; PE, phycoerythrin. Risk factors for severe outcomes following 2009 influenza A (H1N1) infection: a global pooled analysis, PLoS Med, Vol. 8, 2011, e1001053 4 S.D. Message, V. Laza-Stanca, P. Mallia, H.L. Parker, J. Zhu, T. Kebadze, Proc Natl Acad Sci U S A, Vol. 105, 2008, 13562-13567 5...
Phosphoinositides are membrane phospholipids involved in a variety of cellular processes like growth, development, metabolism, and transport. This review focuses on the maintenance of cellular ...homeostasis of phosphatidylinositol 4,5-bisphosphate (PIP
), and phosphatidylinositol 3,4,5-trisphosphate (PIP
). The critical balance of these PIPs is crucial for regulation of neuronal form and function. The activity of PIP
and PIP
can be regulated through kinases, phosphatases, phospholipases and cholesterol microdomains. PIP
and PIP
carry out their functions either indirectly through their effectors activating integral signaling pathways, or through direct regulation of membrane channels, transporters, and cytoskeletal proteins. Any perturbations to the balance between PIP
and PIP
signaling result in neurodevelopmental and neurodegenerative disorders. This review will discuss the upstream modulators and downstream effectors of the PIP
and PIP
signaling, in the context of neuronal health and disease.
Phosphatase and tensin homolog deleted on chromosome 10 (PTEN) protein is a negative regulator of the AKT serine threonine kinases (Protein Kinase B) in the mechanistic target of rapamycin (mTOR) ...signaling pathway. Mutations in PTEN are found in patients with autism, epilepsy, or macrocephaly. In mouse models, Pten-loss results in neuronal hypertrophy, hyperexcitability, seizures, and social deficits. The underlying molecular mechanisms of these phenotypes are not well-delineated. I aimed to disentangle the role of the downstream mTOR complex 1 (mTORC1) and mTOR complex 2 (mTORC2) in orchestrating aberrations in neuronal form and function resulting from Pten-loss. First, we found that the rapamycin-mediated inhibition of mTORC1 in dentate gyrus granule neurons prevented the increase in soma size, migration, spine density and dendritic overgrowth due to lack of Pten. Regulatory-associated protein of mTOR (Raptor) is a scaffolding protein for the formation of themTORC1 complex. Postnatal genetic knockout (KO) of Raptor to disrupt mTORC1 complex formation blocked neuronal hypertrophy observed with Pten-loss. Electrophysiological recordings from cultured neurons revealed that genetic disruption of mTORC1 rescued the increase in excitatory synaptic transmission observed with loss of Pten. Thus, we have identified an essential role for mTORC1 in orchestrating the observed neuronal hypertrophy and synapse formation. Second, as rapamycin-insensitive companion of mTOR (Rictor) is a scaffolding protein for formation of the mTORC2 complex, I found that knocking-out Rictor gene is sufficient to restore normal levels of spine density, and distal dendritic growth. Rictor KO partially rescued somatic hypertrophy, migration deficits, and proximal dendritic branching resulting from Pten-loss. Simultaneous disruption of both mTORC1 and mTORC2 did not result in an additive rescue of somatic hypertrophy, suggesting a role for mTORC1 in the rescue mediated by mTORC2. mTORC2-driven phosphorylation of AKT proteins, a family of kinases upstream of mTORC1, provides one such point of convergence. Disruption of mTORC2 decreased phosphorylation of wildtype AKT3 but did not rescue the morphological effects seen on reconstitution with a phosphomimetic AKT3. Taken together, the data suggest that the rescue of the somatic hypertrophy of Pten KO neurons resulting from Rictor-loss is dependent on the phosphorylation of AKT3. Loss of this phosphorylation downregulates the activity of mTORC1, resulting in a partial restoration of somatic hypertrophy of neurons lacking Pten. Lastly, I found that pharmacologically decreasing the rate of microtubule (MT) polymerization in granule neurons is sufficient to reduce dendritic overgrowth in vivo, as well as to improve the spatial navigation and memory deficits of Pten mutant mice. By teasing apart the molecular signaling underlying neuronal alterations associated with Pten-loss, I have provided mechanistic insight and identified potential new targets for development of therapeutics for autism spectrum disorder.
Hyperactivation of PI3K/PTEN-mTOR signaling during neural development is associated with focal cortical dysplasia (FCD), autism, and epilepsy. mTOR can signal through two major hubs, mTORC1 and ...mTORC2, both of which are hyperactive following PTEN loss of function (LOF). Here, we tested the hypothesis that genetic inactivation of the mTORC2 complex via deletion of
is sufficient to rescue morphologic and electrophysiological abnormalities in the dentate gyrus caused by PTEN loss, as well as generalized seizures. An established, early postnatal mouse model of PTEN loss in male and female mice showed spontaneous seizures that were not prevented by mTORC2 inactivation. This lack of rescue occurred despite the normalization or amelioration of many morphologic and electrophysiological phenotypes. However, increased excitatory connectivity proximal to dentate gyrus granule neuron somas was not normalized by mTORC2 inactivation. Further studies demonstrated that, although mTORC2 inactivation largely rescued the dendritic arbor overgrowth caused by PTEN LOF, it increased synaptic strength and caused additional impairments of presynaptic function. These results suggest that a constrained increase in excitatory connectivity and co-occurring synaptic dysfunction is sufficient to generate seizures downstream of PTEN LOF, even in the absence of characteristic changes in morphologic properties.
Homozygous deletion of the
gene in neuronal subpopulations in the mouse serves as a valuable model of epilepsy caused by mTOR hyperactivation. To better understand the physiological mechanisms downstream of
loss that cause epilepsy, as well as the therapeutic potential of targeted gene therapies, we tested whether genetic inactivation of the mTORC2 complex could improve the cellular, synaptic, and
effects of
loss in the dentate gyrus. We found that mTORC2 inhibition improved or rescued all morphologic effects of
loss in the dentate gyrus, but synaptic changes and seizures persisted. These data suggest that synaptic dysfunction can drive epilepsy caused by hyperactivation of PI3K/PTEN-mTOR, and that future therapies should focus on this mechanistic link.
Background Asthma is a heterogeneous disease in which there is a differential response to asthma treatments. This heterogeneity needs to be evaluated so that a personalized management approach can be ...provided. Objectives We stratified patients with moderate-to-severe asthma based on clinicophysiologic parameters and performed an omics analysis of sputum. Methods Partition-around-medoids clustering was applied to a training set of 266 asthmatic participants from the European Unbiased Biomarkers for the Prediction of Respiratory Diseases Outcomes (U-BIOPRED) adult cohort using 8 prespecified clinic-physiologic variables. This was repeated in a separate validation set of 152 asthmatic patients. The clusters were compared based on sputum proteomics and transcriptomics data. Results Four reproducible and stable clusters of asthmatic patients were identified. The training set cluster T1 consists of patients with well-controlled moderate-to-severe asthma, whereas cluster T2 is a group of patients with late-onset severe asthma with a history of smoking and chronic airflow obstruction. Cluster T3 is similar to cluster T2 in terms of chronic airflow obstruction but is composed of nonsmokers. Cluster T4 is predominantly composed of obese female patients with uncontrolled severe asthma with increased exacerbations but with normal lung function. The validation set exhibited similar clusters, demonstrating reproducibility of the classification. There were significant differences in sputum proteomics and transcriptomics between the clusters. The severe asthma clusters (T2, T3, and T4) had higher sputum eosinophilia than cluster T1, with no differences in sputum neutrophil counts and exhaled nitric oxide and serum IgE levels. Conclusion Clustering based on clinicophysiologic parameters yielded 4 stable and reproducible clusters that associate with different pathobiological pathways.
Genetic studies of hippocampal granule neuron development have been used to elucidate cellular functions of Pten and Fmr1. While mutations in each gene cause neurodevelopmental disorders such as ...autism and fragile X syndrome, how Pten and Fmr1 function alone or together during normal development is not known. Moreover, Pten mRNA is bound by the fragile X mental retardation protein (FMRP) RNA binding protein, but how this physical interaction impinges on phosphatase and tensin homolog protein (PTEN) expression is not known. To understand the interaction of PTEN and FMRP, we investigated the dentate gyrus granule neuron development in Pten and Fmr1 knockout (KO) mice. Interestingly, heterozygosity of Pten restored Fmr1 KO cellular phenotypes, including dendritic arborization, and spine density, while PTEN protein expression was significantly increased in Fmr1 KO animals. However, complete deletion of both Pten and Fmr1 resulted in a dramatic increase in dendritic length, spine density, and spine length. In addition, overexpression of PTEN in Fmr1 KO Pten heterozygous background reduced dendritic length, arborization, spine density, and spine length including pS6 levels. Our findings suggest that PTEN levels are negatively regulated by FMRP, and some Fmr1 KO phenotypes are caused by dysregulation of PTEN protein. These observations provide evidence for the genetic interaction of PTEN and FMRP and a possible mechanistic basis for the pathogenesis of Fmr1-related fragile X neurodevelopmental disorders.
Phosphatase and tensin homolog (PTEN) is a major negative regulator of the phosphatidylinositol-3-kinase (PI3K)/Akt/mechanistic target of rapamycin (mTOR) pathway. Loss-of-function mutations in
have ...been found in a subset of patients with macrocephaly and autism spectrum disorder (ASD). PTEN loss in neurons leads to somal hypertrophy, aberrant migration, dendritic overgrowth, increased spine density, and hyperactivity of neuronal circuits. These neuronal overgrowth phenotypes are present on
knock-out (KO) and reconstitution with autism-associated point mutations. The mechanism underlying dendritic overgrowth in
deficient neurons is unclear. In this study, we examined how
loss impacts microtubule (MT) dynamics in both sexes using retroviral infection and transfection strategies to manipulate PTEN expression and tag the plus-end MT binding protein, end-binding protein 3 (EB3). We found
KO neurons sprout more new processes over time compared with wild-type (WT) neurons. We also found an increase in MT polymerization rate in
KO dendritic growth cones. Reducing MT polymerization rate to the WT level was sufficient to reduce dendritic overgrowth in
KO neurons
and
Finally, we found that rescue of dendritic overgrowth via inhibition of MT polymerization was sufficient to improve the performance of
KO mice in a spatial memory task. Taken together, our data suggests that one factor underlying PTEN loss dependent dendritic overgrowth is increased MT polymerization. This opens the possibility for an intersectional approach targeting MT polymerization and mTOR with low doses of inhibitors to achieve therapeutic gains with minimal side effects in pathologies associated with loss of neuronal PTEN function.
Loss of
function because of genetic deletion or expression of mutations associated with autism spectrum disorder (ASD), results in overgrowth of neurons including increased total dendritic length and branching. We have discovered that this overgrowth is accompanied by increased rate of microtubule (MT) polymerization. The increased polymerization rate is insensitive to acute inhibition of mechanistic target of rapamycin (mTOR)C1 or protein synthesis. Direct pharmacological inhibition of MT polymerization can slow the polymerization rate in
knock-out (KO) neurons to rates seen in wild-type (WT) neurons. Correction of the MT polymerization rate rescues increased total dendritic arborization and spatial memory. Our studies suggest that phosphatase and tensin homolog (PTEN) inhibits dendritic growth through parallel regulation of protein synthesis and cytoskeletal polymerization.
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