An Activity-Guided Map of Electrophile-Cysteine Interactions in Primary Human T Cells.

Electrophilic compounds originating from nature or chemical synthesis have profound effects on immune cells. These compounds are thought to act by cysteine modification to alter the functions of immune-relevant proteins; however, our understanding of electrophile-sensitive cysteines in the human immune proteome remains limited. Here, we present a global map of cysteines in primary human T cells that are susceptible to covalent modification by electrophilic small molecules. More than 3,000 covalently liganded cysteines were found on functionally and structurally diverse proteins, including many that play fundamental roles in immunology. We further show that electrophilic compounds can impair T cell activation by distinct mechanisms involving the direct functional perturbation and/or degradation of proteins. Our findings reveal a rich content of ligandable cysteines in human T cells and point to electrophilic small molecules as a fertile source for chemical probes and ultimately therapeutics that modulate immunological processes and their associated disorders.

Affinity-Guided Oxime Chemistry for Selective Protein Acylation in Live Tissue Systems.

Catalyst-mediated protein modification is a powerful approach for the imaging and engineering of natural proteins. We have previously developed affinity-guided 4-dimethylaminopyridine (AGD) chemistry as an efficient protein modification method using a catalytic acyl transfer reaction. However, because of the high electrophilicity of the thioester acyl donor molecule, AGD chemistry suffers from nonspecific reactions to proteins other than the target protein in crude biological environments, such as cell lysates, live cells, and tissue samples. To overcome this shortcoming, we here report a new acyl donor/organocatalyst system that allows more specific and efficient protein modification. In this method, a highly nucleophilic pyridinium oxime (PyOx) catalyst is conjugated to a ligand specific to the target protein. The ligand-tethered PyOx selectively binds to the target protein and facilitates the acyl transfer reaction of a mild electrophilic N-acyl-N-alkylsulfonamide acyl donor on the protein surface. We demonstrated that the new catalytic system, called AGOX (affinity-guided oxime) chemistry, can modify target proteins, both in test tubes and cell lysates, more selectively and efficiently than AGD chemistry. Low-background fluorescence labeling of the endogenous cell-membrane proteins, carbonic anhydrase XII and the folate receptor, in live cells allowed for the precise quantification of diffusion coefficients in the protein's native environment. Furthermore, the excellent biocompatibility and bioorthogonality of AGOX chemistry were demonstrated by the selective labeling of an endogenous neurotransmitter receptor in mouse brain slices, which are highly complicated tissue samples.

Identification of protein aggregates in the aging vertebrate brain with prion-like and phase-separation properties.

Protein aggregation, which can sometimes spread in a prion-like manner, is a hallmark of neurodegenerative diseases. However, whether prion-like aggregates form during normal brain aging remains unknown. Here, we use quantitative proteomics in the African turquoise killifish to identify protein aggregates that accumulate in old vertebrate brains. These aggregates are enriched for prion-like RNA-binding proteins, notably the ATP-dependent RNA helicase DDX5. We validate that DDX5 forms aggregate-like puncta in the brains of old killifish and mice. Interestingly, DDX5's prion-like domain allows these aggregates to propagate across many generations in yeast. In vitro, DDX5 phase separates into condensates. Mutations that abolish DDX5 prion propagation also impair the protein's ability to phase separate. DDX5 condensates exhibit enhanced enzymatic activity, but they can mature into inactive, solid aggregates. Our findings suggest that protein aggregates with prion-like properties form during normal brain aging, which could have implications for the age-dependency of cognitive decline.

An appropriate treatment interval does not affect the prognosis of patients with breast Cancer.

Purpose: Major public health emergencies may lead to delays or alterations in the treatment of patients with breast cancer at each stage of diagnosis and treatment. How much do these delays and treatment changes affect treatment outcomes in patients with breast cancer? Methods: This review summarized relevant research in the past three decades and identified the effect of delayed treatment on the prognosis of patients with breast cancer in terms of seeking medical treatment, neoadjuvant treatment, surgery, postoperative chemotherapy, radiotherapy, and targeted therapies. Results: Delay in seeking medical help for ≥12 weeks affected the prognosis. Surgical treatment within 4 weeks of diagnosis did not affect patient prognosis. Starting neoadjuvant chemotherapy within 8 weeks after diagnosis, receiving surgical treatment at 8 weeks or less after the completion of neoadjuvant chemotherapy, and receiving radiotherapy 8 weeks after surgery did not affect patient prognosis. Delayed chemotherapy did not increase the risk of relapse in patients with luminal A breast cancer. Every 4 weeks of delay in the start of postoperative chemotherapy in patients with luminal B, triple-negative, or HER2-positive breast cancer treated with trastuzumab will adversely affect the prognosis. Targeted treatment delays in patients with HER2-positive breast cancer should not exceed 60 days after surgery or 4 months after diagnosis. Radiotherapy within 8 weeks after surgery did not increase the risk of recurrence in patients with early breast cancer who were not undergoing adjuvant chemotherapy. Conclusion: Different treatments have different time sensitivities, and the careful evaluation and management of these delays will be helpful in minimizing the negative effects on patients.