Biology subjects

Webster, R.

Publications and source records attributed to Webster, R..

4 recordsLinked to original sources

bioRxiv2026

Role of TbTim17 in mitochondrial stress tolerance in Trypanosoma brucei

Mitochondrial protein translocases Tim17 and Tim23 play important roles in stress-response pathways via activating the transcription factors, ATFS-1, to maintain organellar homeostasis. Trypanosoma brucei, a divergent eukaryote and the infectious agent for African trypanosomiasis, lacks ATFs but possesses TbTim17, an essential component of the TIM complex in mitochondria. However, it has not been investigated whether TbTim17 plays a role in the mitochondrial stress response. Here, we show that depletion of TbTim17 increased T. brucei tolerance to paraquat, increasing the EC50 by 3- to 4-fold compared with the wild type. Subsequent analysis revealed that increased levels of mitochondrial reactive oxygen species resulting from TbTim17 knockdown upregulate mitochondrial superoxide dismutase, thereby preadapting cells to resist paraquat-induced oxidative stress. This is supported by the finding that treating cells with N-acetyl cysteine during TbTim17 RNAi induction reduced the EC50 of paraquat to wild-type levels. TbTim17 knockdown also increased tolerance of T. brucei to heat stress. Either heat or oxidative stress did not increase mitochondrial heat shock protein 70 or Bip levels in the ER in T. brucei; instead, they moderately increased TbTim17 levels, which can replenish mitochondrial proteomes. Furthermore, TbTim17 knockdown caused a significant reduction in SL RNA and a 2- to 5-fold increase in tSNAP42 transcript levels, suggesting that mitochondrial stress is linked to the ER stress response pathway in T. brucei. Together, these results show that the mitochondrial stress response is primarily mediated by antioxidant defense mechanisms and that TbTim17 plays a protective role in mitochondria under stress in T. brucei. ImportanceTrypanosoma brucei, a parasitic protozoan, is the infectious agent of a deadly disease in humans and livestock known as African trypanosomiasis. TbTim17 is the major component of the TbTIM complex that imports hundreds of nuclear-encoded proteins into the mitochondrial matrix and inner membrane. Here, we show that depletion of TbTim17 induces oxidative stress, which upregulates mitochondrial antioxidant defense mechanisms to mitigate this stress. Subsequently, the SLS response is induced to eliminate the defective parasite from the population. In the wild-type parasite, TbTim17 levels are increased in mitochondria under oxidative and heat stresses, likely to replenish damaged mitochondrial proteomes. This is unlike in other eukaryotes, where oxidative stress degrades Tim17 to induce mitochondrial stress response. Understanding the mitochondrial stress response in T. brucei and the role of mitochondrial protein translocases in this process is critical for elucidating the mechanisms of adaptation to environmental stresses and drug resistance in this parasite.

Cell Biology↗

bioRxiv2024

Structures of the human adult muscle-type nicotinic receptor in resting and desensitised states

Muscle-type nicotinic acetylcholine receptor (AChR) is the key signalling molecule in neuromuscular junctions. Here we present the structures of full-length human adult receptors in complex with Fab35 in -bungarotoxin (BuTx)-bound resting states, and in acetylcholine (ACh)-bound desensitised states. In addition to identifying the conformational changes during recovery from desensitisation, we also used electrophysiology to probe the effects of eight previously unstudied AChR genetic variants found in congenital myasthenic syndrome (CMS) patients, revealing they cause either slow- or fast-channel CMS characterised by prolonged or abbreviated ion channel bursts. The combined kinetic and structural data offer a better understanding of both AChR state transition and the pathogenic mechanisms of disease variants.

neuroscience↗

bioRxiv2024

The catheterized bladder environment induces dysregulation of macrophage polarization exacerbating bacterial UTI

Urinary catheterization causes bladder damage, predisposing hosts to catheter-associated urinary tract infections (CAUTIs). CAUTI pathogenesis is mediated by bladder damage-induced inflammation, resulting in accumulation and deposition of the blood-clotting protein fibrinogen (Fg) and its matrix form fibrin, which are exploited by uropathogens as biofilm platforms to establish infection. Catheter-induced inflammation also results in robust immune cell recruitment, including macrophages (M{phi}s). A fundamental knowledge gap is understanding the mechanisms by which the catheterized-bladder environment suppresses the M{phi} antimicrobial response, allowing uropathogen persistence. Here, we found that Fg and fibrin differentially modulate M1 and M2 M{phi} polarization, respectively. We unveiled that fibrin accumulation in catheterized mice induced an anti-inflammatory M2-like M{phi} phenotype, correlating with pathogen persistence. Even GM-CSF treatment of wildtype mice to promote M1 polarization was not sufficient to reduce bacterial burden and dissemination, indicating that the catheterized-bladder environment provides mixed signals, dysregulating M{phi} polarization, hindering its antimicrobial response against uropathogens.

immunology↗

bioRxiv2020

Cell size homeostasis is maintained by a circuitry involving a CDK4-determined target size that programs the cell size-dependent activation of p38

While molecules that promote the growth of animal cells have been identified, the following question remains: How are growth promoting pathways regulated to specify a characteristic size for each of the different cell types? In 1975, Hartwell and Nurse suggested that in eukaryotes, cell size is determined by size checkpoints - mechanisms that restrict cell cycle progression from cells that are smaller than their target size. Curiously, such checkpoint mechanisms imply a conceptual distinction between a cells actual size and cells target size. In the present study, we materialize this conceptual distinction by describing experimental assays that discriminately quantify a cells target size value. With these assays, we show that a cells size and target size are distinct phenotypes that are subject to different upstream regulators. While mTORC1 promotes growth in cell size, our data suggests that a cells target size value is regulated by other pathways including FGFR3, ROCK2, and CDK4. For example, while rapamycin (an mTORC1 inhibitor) decreases cell size, rapamycin does not change the target size that is required for the G1/S transition. The CDK4/Rb pathway has been previously proposed as a putative regulator of target size. Yet, in lacking experimental means that discriminate perturbations of cell growth from perturbations that reprogram target size, such claims on target size were not validated. To investigate the functions of CDK4 in target size determination, we used genetic and chemical means to dial higher and lower levels of CDK4 activity. These measurements identified functions of CDK4 on target size that are distinct from other G1 CDKs. Using C. elegans, we further demonstrate that these influences of CDK4 on size determination function in vivo. Finally, we propose a model whereby mTORC1, p38, and CDK4 cooperate in a manner that is analogous to the function of a thermostat. While mTORC1 promotes cellular growth as prompted by p38, CDK4 is analogous to the thermostat dial that sets the critical target size associated with cell size homeostasis.

cell biology↗