Biology subjects

Turner, S. R.

Publications and source records attributed to Turner, S. R..

5 recordsLinked to original sources

bioRxiv2026

Leveraging HIV-Specific CAR T Cells and Rapamycin Treatment in Kick-and-Kill HIV Cure Approaches

HIV is not cured with currently available combination antiretroviral therapy (ART) alone in large part because the virus establishes virologic latency in long lived CD4+ cells. To eliminate this latent reservoir, as required for HIV eradication, latency reversing agents (LRAs) are being developed to force HIV out of latency and induce infected cells to express viral proteins leading to their clearance, in a so-called "Kick-and-Kill" approach. This strategy relies on the immune system to clear the productively-infected cells and is thus limited by HIV immune evasion and the immunological exhaustion that occurs during HIV infection. To counter these limitations and augment an LRA-mediated HIV cure approach, we report herein the utility of HIV-specific truncated CD4-based D1D2 CAR T cells combined with LRA treatment and the mTORC1 inhibitor rapamycin to reduce immune exhaustion and specifically target and kill LRA-stimulated HIV infected cells. We demonstrate that rapamycin does not prevent HIV latency reversal via multiple classes of LRAs in several in vitro models, suggesting that it is compatible with cure approaches utilizing these LRAs. Additionally, rapamycin does not inhibit the early T cell activation (CD69 upregulation) in primary T cells that occurs during protein kinase C (PKC) modulator-mediated HIV latency reversal. Furthermore, in vitro chronically exhausted CAR T cells were found to have a higher frequency of terminally exhausted PD-1+Tim-3+ and CD69+PD-1+ cells when compared to CAR T cells that were cultured under the same conditions in the presence of rapamycin, validating the use of the mTORC1 inhibitor rapamycin to prevent immune exhaustion of CAR T cells. Finally, we found that latently-infected cells that were stimulated to express HIV proteins using a designed, synthetic PKC modulator LRA (SUW133) were efficiently recognized and killed by CAR T cells. Overall, these data demonstrate the compatibility of immune rejuvenation using rapamycin with HIV reservoir depletion using LRAs and CAR T cells. This combination therapy strategy represents a promising approach to more effectively target the latent reservoir in HIV cure approaches.

immunology↗

bioRxiv2024

Cellulose synthesis in Arabidopsis by heterologous expression of lower plants CESA proteins

Cellulose is the most abundant component of plant cell walls where it plays a pivotal role in regulating plant cell size and shape. In addition, as a component of the woody secondary cell walls, cellulose represents an abundant renewable resource to produce materials and chemicals. In higher plants, cellulose is synthesised at the plasma membrane by a hexameric protein complex, known as the rosette, that is able to synthesise 18 glucose chains that bond together to form a microfibril. While this rosette structure is highly conserved, significant variation exists in the structure and physical properties of cellulose found in different cell types and synthesised by different species. In this study, we surveyed the ability of the catalytic subunits of the cellulose synthase complex (CESA proteins) from a range of lower plant species to synthesise cellulose in the Arabidopsis secondary cell walls. Several lower plant CESA proteins are able to function in higher plants in conjunction Arabidopsis CESAs. Additionally, two moss CESA proteins synthesised cellulose in absence of Arabidopsis CESAs but with reduced crystallinity, indicating that it is the structure of CESA proteins themselves and not the cellular environment that determines the properties of the cellulose synthesised.

plant biology↗

bioRxiv2022

Death-associated protein kinase 3 (DAPK3) regulates the myogenic reactivity of cerebral arterioles

The vascular smooth muscle (VSM) of resistance blood vessels displays intrinsic autoregulatory responses to increased intraluminal pressure, the myogenic response. In the brain, the myogenic responses of cerebral arterioles are critical to homeostatic blood flow regulation. Here we provide the first evidence to link the death-associated protein kinase 3 (DAPK3) to the myogenic response of rat and human cerebral arterioles. DAPK3 is a Ser/Thr kinase involved in Ca2+- sensitization mechanisms of VSM contraction. Ex vivo administration of a specific DAPK3 inhibitor (i.e., HS38) could attenuate vessel constrictions invoked by serotonin as well as intraluminal pressure elevation. The HS38-dependent dilation was not associated with any change in myosin light chain (LC20) phosphorylation. The results suggest that DAPK3 does not regulate Ca2+ sensitization pathways during the myogenic response of cerebral vessels but rather operates to control the actin cytoskeleton. Finally, a slow return of myogenic tone was observed during the sustained exposure of cerebral arterioles to a suite of DAPK3 inhibitors. Recovery of tone was associated with greater LC20 phosphorylation that suggests intrinsic signaling compensation in response to attenuation of DAPK3 activity. The translational importance of DAPK3 to the human cerebral vasculature was noted, with robust expression of the protein kinase and significant HS38-dependent attenuation of myogenic reactivity found for human pial vessels.

physiology↗

bioRxiv2020

Enhanced diastolic dysfunction but preserved systolic function after acute pressure overload in the absence of smoothelin-like 1 protein

AimsSmoothelin-like 1 (SMTNL1), a protein kinase A/G target protein, modulates the activity and expression of myosin light chain phosphatase and thus plays an important role in regulating vasoconstriction. Increased myogenic reactivity of resistance arterioles is associated with SMTNL1 silencing, and elevated baseline vascular tone is increasingly recognized as a risk factor for development of hypertension and chronic congestive heart failure. Hence, in this study we assessed cardiac function in SMTNL1 knockout mice with and without accompanying acute cardiac stress (i.e., pressure overload by transverse aortic constriction). Methods and ResultsMale and female, global Smtnl1 knockout (KO) & wild-type (WT) mice were assessed at 10 weeks of age by echocardiography and electrocardiography to define baseline cardiac function. Gross dissection revealed distinct cardiac morphology only in male mice; hearts from KO animals were significantly smaller than WT littermates but the proportion of heart mass taken up by LV was greater. Non-invasive analyses of KO mice showed reduced resting heart rate with improved ejection fraction and fractional shortening as well as elevated aortic and pulmonary flow velocities relative to their WT counterparts, but only in the male cohort. We further investigated the impact of acute pressure overload on cardiac morphometry and hemodynamics in the absence of SMTNL1 in male cohort using echocardiography and pressure-volume (PV) loop measurements. Interestingly, PV loop analysis revealed diastolic dysfunction with significantly increased end diastolic pressure and LV relaxation time along with a steeper end diastolic pressure-volume relationship an indicator of stiffer heart, in the KO group when compared to WT Sham-operated group. Sham KO mice also showed elevated arterial elastance and total peripheral resistance. With acute pressure overload, systolic function was preserved, but diastolic dysfunction was exacerbated in KO mice with higher E/E ratio and myocardial performance index along with a prolonged isovolumetric relaxation time relative to the aortic-banded WT group. ConclusionTaken together, the findings support a novel, sex-dimorphic role for SMTNL1 in modulating cardiac structure and diastolic function. Significantly, impairment of diastolic function following pressure overload in young animals lacking SMTNL1 is mainly driven by increased systemic vascular resistance, which mimics the clinical pathophysiology of heart failure with preserved ejection fraction (HFpEF). Translational PerspectiveHeart failure with preserved ejection fraction (HFpEF) is characterized by the impairment of diastolic function and accounts for half of all heart failure cases. Unfortunately, there is as yet no proven therapy available for these patients as the pathophysiology is complicated with the presence of multiple comorbidities, microvascular dysfunction and a lack of an ideal animal model. The phenotype of Smtnl1 global deletion male mice exhibits intriguing similarities to HFpEF, with elevated microvascular resistance driving diastolic dysfunction and LV remodeling. As such the SMTNL1 KO mouse represents a novel pre-clinical model to study the molecular etiology of HFpEF.

physiology↗

bioRxiv2020

An atlas of Arabidopsis protein S-Acylation reveals its widespread role in plant cell organisation of and function.

S-acylation is the addition of a fatty acid to a cysteine residue of a protein. While this modification may profoundly alter protein behaviour, its effects on the function of plant proteins remains poorly characterised, largely as a result to the lack of basic information regarding which proteins are S-acylated and where in the proteins the modification occurs. In order to address this gap in our knowledge, we have performed a comprehensive analysis of plant protein S-acylation from 6 separate tissues. In our highest confidence group, we identified 5185 cysteines modified by S-acylation, which were located in 4891 unique peptides from 2643 different proteins. This represents around 9% of the entire Arabidopsis proteome and suggests an important role for S-acylation in many essential cellular functions including trafficking, signalling and metabolism. To illustrate the potential of this dataset, we focus on cellulose synthesis and confirm for the first time the S-acylation of all proteins known to be involved in cellulose synthesis and trafficking of the cellulose synthase complex. In the secondary cell walls, cellulose synthesis requires three different catalytic subunits (CESA4, CESA7 and CESA8) that all exhibit striking sequence similarity. While all three proteins have been widely predicted to possess a RING-type zinc finger at their N-terminus, for CESA4 and CESA8, we find evidence for S-acylation of cysteines in this region that is incompatible with any role in coordinating metal ions. We show that while CESA7 may possess a RING type domain, the same region of CESA4 and CESA8 appear to have evolved a very different structure. Together, the data suggests this study represents an atlas of S-acylation in Arabidopsis that will facilitate the broader study of this elusive post-translational modification in plants as well as demonstrates the importance of undertaking further work in this area.

plant biology↗