Alterations in cAMP signaling are thought to contribute to neurocognitive and neuropsychiatric disorders. Members of the cAMPspecific phosphodiesterase 4 (PDE4) family, which contains Ͼ25 different isoforms, play a key role in determining spatial cAMP degradation so as to orchestrate compartmentalized cAMP signaling in cells. Each isoform binds to a different set of protein complexes through its unique N-terminal domain, thereby leading to targeted degradation of cAMP in specific intracellular compartments. However, the functional role of specific compartmentalized PDE4 isoforms has not been examined in vivo. Here, we show that increasing protein levels of the PDE4A5 isoform in mouse hippocampal excitatory neurons impairs a long-lasting form of hippocampal synaptic plasticity and attenuates hippocampus-dependent long-term memories without affecting anxiety. In contrast, viral expression of a truncated version of PDE4A5, which lacks the unique N-terminal targeting domain, does not affect long-term memory. Further, overexpression of the PDE4A1 isoform, which targets a different subset of signalosomes, leaves memory undisturbed. Fluorescence resonance energy transfer sensorbased cAMP measurements reveal that the full-length PDE4A5, in contrast to the truncated form, hampers forskolin-mediated increases in neuronal cAMP levels. Our study indicates that the unique N-terminal localization domain of PDE4A5 is essential for the targeting of specific cAMP-dependent signaling underlying synaptic plasticity and memory. The development of compounds to disrupt the compartmentalization of individual PDE4 isoforms by targeting their unique N-terminal domains may provide a fruitful approach to prevent cognitive deficits in neuropsychiatric and neurocognitive disorders that are associated with alterations in cAMP signaling.

During pre-synaptic embryonic development, neuronal processes traverse short distances to reach their targets via growth cone. Over time, neuronal somata are separated from their axon terminals due to skeletal growth of the enlarging organism (Weiss 1941;Gray, Hukkanen et al. 1992). This mechanotransduction induces a secondary mode of neuronal growth capable of accommodating continual elongation of the axon (Bray 1984;Heidemann and Buxbaum 1994;Heidemann, Lamoureux et al. 1995;Pfister, Iwata et al. 2004).Axon Stretch Growth (ASG) is conceivably a central factor in the maturation of short embryonic processes into the long nerves and white matter tracts characteristic of the adult nervous system. To study ASG in vitro, we engineered bioreactors to apply tension to the short axonal processes of neuronal cultures (Loverde, Ozoka et al. 2011). Here, we detail the methods we use to prepare bioreactors and conduct ASG. First, within each stretching lane of the bioreactor, neurons are plated upon a micro-manipulated towing substrate. Next, neurons regenerate their axonal processes, via growth cone extension, onto a stationary substrate. Finally, stretch growth is performed by towing the plated cell bodies away from the axon terminals adhered to the stationary substrate; recapitulating skeletal growth after growth cone extension.Previous work has shown that ASG of embryonic rat dorsal root ganglia neurons are capable of unprecedented growth rates up to 10mm/day, reaching lengths of up to 10cm; while concurrently resulting in increased axonal diameters (Smith, Wolf et al. 2001;Pfister, Iwata et al. 2004;Pfister, Bonislawski et al. 2006;Pfister, Iwata et al. 2006;Smith 2009). This is in dramatic contrast to regenerative growth cone extension (in absence of mechanical stimuli) where growth rates average 1mm/day with successful regeneration limited to lengths of less than 3cm (Fu and Gordon 1997;Pfister, Gordon et al. 2011). Accordingly, further study of ASG may help to reveal dysregulated growth mechanisms that limit regeneration in the absence of mechanical stimuli.

The ability to predict responsiveness to drugs in individual patients is limited. We hypothesized that integrating molecular information from databases would yield predictions that could be experimentally tested to develop transcriptomic signatures for specific drugs. We analyzed lung adenocarcinoma patient data from The Cancer Genome Atlas and identified a subset of patients in which xanthine dehydrogenase (XDH) expression correlated with decreased survival. We tested allopurinol, an FDA‐approved drug that inhibits XDH, on human non‐small‐cell lung cancer (NSCLC) cell lines obtained from the Broad Institute Cancer Cell Line Encyclopedia and identified sensitive and resistant cell lines. We utilized the transcriptomic profiles of these cell lines to identify six‐gene signatures for allopurinol‐sensitive and allopurinol‐resistant cell lines. Transcriptomic networks identified JAK2 as an additional target in allopurinol‐resistant lines. Treatment of resistant cell lines with allopurinol and CEP‐33779 (a JAK2 inhibitor) resulted in cell death. The effectiveness of allopurinol alone or allopurinol and CEP‐33779 was verified in vivo using tumor formation in NCR‐nude mice. We utilized the six‐gene signatures to predict five additional allopurinol‐sensitive NSCLC cell lines and four allopurinol‐resistant cell lines susceptible to combination therapy. We searched the transcriptomic data from a library of patient‐derived NSCLC tumors from the Jackson Laboratory to identify tumors that would be predicted to be sensitive to allopurinol or allopurinol + CEP‐33779 treatment. Patient‐derived tumors showed the predicted drug sensitivity in vivo . These data indicate that we can use integrated molecular information from cancer databases to predict drug responsiveness in individual patients and thus enable precision medicine.

Whole cell responses involve multiple subcellular processes (SCPs). To understand how balance between SCPs controls the dynamics of whole cell responses we studied neurite outgrowth in rat primary cortical neurons in culture. We used a combination of dynamical models and experiments to understand the conditions that permitted growth at a specified velocity and when aberrant growth could lead to the formation of dystrophic bulbs. We hypothesized that dystrophic bulb formation is due to quantitative imbalances between SCPs. Simulations predict redundancies between lower level sibling SCPs within each type of high level SCP. In contrast, higher level SCPs, such as vesicle transport and exocytosis or microtubule growth characteristic of each type need to be strictly coordinated with each other and imbalances result in stalling of neurite outgrowth. From these simulations, we predicted the effect of changing the activities of SCPs involved in vesicle exocytosis or microtubule growth could lead to formation of dystrophic bulbs. siRNA ablation experiments verified these predictions. We conclude that whole cell dynamics requires balance between the higher-level SCPs involved and imbalances can terminate whole cell responses such as neurite outgrowth.

SARS-CoV-2 infects multiple organs, including the heart. Analyses of hospitalized patients show that a substantial number without prior indication of heart disease or comorbidities show significant injury to heart tissue, assessed by increased levels of troponin in blood.

COVID-19 affects multiple organs. Clinical data from the Mount Sinai Health System shows that substantial numbers of COVID-19 patients without prior heart disease develop cardiac dysfunction. How COVID-19 patients develop cardiac disease is not known. We integrate cell biological and physiological analyses of human cardiomyocytes differentiated from human induced pluripotent stem cells (hiPSCs) infected with SARS-CoV-2 in the presence of interleukins, with clinical findings, to investigate plausible mechanisms of cardiac disease in COVID-19 patients. We infected hiPSC-derived cardiomyocytes, from healthy human subjects, with SARS-CoV-2 in the absence and presence of interleukins. We find that interleukin treatment and infection results in disorganization of myofibrils, extracellular release of troponin-I, and reduced and erratic beating. Although interleukins do not increase the extent, they increase the severity of viral infection of cardiomyocytes resulting in cessation of beating. Clinical data from hospitalized patients from the Mount Sinai Health system show that a significant portion of COVID-19 patients without prior history of heart disease, have elevated troponin and interleukin levels. A substantial subset of these patients showed reduced left ventricular function by echocardiography. Our laboratory observations, combined with the clinical data, indicate that direct effects on cardiomyocytes by interleukins and SARS-CoV-2 infection can underlie the heart disease in COVID-19 patients.One Sentence SummaryCardiomyocytes derived from human induced pluripotent stem cells treated with interleukins and infected with SARS- CoV- 2 in cultures, show increased release of troponin, disorganization of myofibrils, and changes in beating mirroring specific pathologies in some COVID-19 patients.

Dopamine, a key striatal neuromodulator, increases synaptic strength by promoting surface insertion and/or retention of AMPA receptors (AMPARs). This process is mediated by the phosphorylation of the GluA1 subunit of AMPAR by cyclic nucleotide-dependent kinases, making cyclic nucleotide phosphodiesterases (PDEs) potential regulators of synaptic strength. In this study, we examined the role of phosphodiesterase 2 (PDE2), a medium spiny neuron-enriched and cGMP-activated PDE, in AMPAR trafficking. We found that inhibiting PDE2 resulted in enhancement of dopamine-induced surface GluA1 expression in dopamine receptor 1-expressing medium spiny neurons. Using pharmacological and genetic approaches, we found that inhibition of PDE1 resulted in a decrease in surface AMPAR levels because of the allosteric activation of PDE2. The cross-regulation of PDE1 and PDE2 activities results in counterintuitive control of surface AMPAR expression, making it possible to regulate the directionality and magnitude of AMPAR trafficking.

27Whole cell responses involve multiple subcellular processes (SCPs). To understand how balance 28 between SCPs controls the dynamics of whole cell responses we studied neurite outgrowth in rat 29 primary cortical neurons in culture. We used a combination of dynamical models and experiments 30 to understand the conditions that permitted growth at a specified velocity and when aberrant 31 growth could lead to the formation of dystrophic bulbs. We hypothesized that dystrophic bulb 32 formation is due to quantitative imbalances between SCPs. Simulations predict redundancies 33 between lower level sibling SCPs within each type of high level SCP. In contrast, higher level 34 SCPs, such as vesicle transport and exocytosis or microtubule growth characteristic of each type 35 need to be strictly coordinated with each other and imbalances result in stalling of neurite 36 outgrowth. From these simulations, we predicted the effect of changing the activities of SCPs 37 involved in vesicle exocytosis or microtubule growth could lead to formation of dystrophic bulbs. 38siRNA ablation experiments verified these predictions. We conclude that whole cell dynamics 39 requires balance between the higher-level SCPs involved and imbalances can terminate whole cell 40 responses such as neurite outgrowth. 41 42 neurite outgrowth, multi compartment model, ordinary differential equations 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 Neurite outgrowth is an early event that changes the state of neurons (Arimura and Kaibuchi, 2007) 58 and allows neurons to develop axons and dendritic trees that connect to other neurons and become 59 electrically active. When there is nerve injury and the axons are severed, the process of 60 regeneration which is similar to neurite outgrowth often fails, resulting in the formation of bulbs 61 at the ends of regenerating axons. These bulbs are called dystrophic bulbs (Hill, 2017). Although 62 some of the cellular pathways involved in axonal regeneration, failure and formation of the 63 dystrophic bulbs are known (Blanquie and Bradke, 2018; Filious and Schwab, 2018) the 64 mechanisms by which these lead to failure are not well understood. In this study we have used an 65 integrated computational and experimental approach to understand the origins of dystrophic bulbs 66 and more generally the conditions under which whole cell responses such as neurite outgrowth can 67 be maintained. The central question we asked in this study is whether quantitative imbalances 68 between subcellular processes can lead to the formation of dystrophic bulbs. 69 Whole cell responses that involve, both morphological and physiological changes, are complex 70 because they can engage many subcellular processes (SCPs) that are together responsible for whole 71 cell functions. SCPs include biochemical pathways and cell biological processes involved the 72 functions of different organelles. Coordinated changes in the activities of SCPs often lead to a 73 change in cell state, such as moving to a more differentiated phenotype. Deta...