Polarized Proteins in Endothelium and Their Contribution to Function.

Protein localization in endothelial cells is tightly regulated to create distinct signaling domains within their tight spatial restrictions including luminal membranes, abluminal membranes, and interendothelial junctions, as well as caveolae and calcium signaling domains. Protein localization in endothelial cells is also determined in part by the vascular bed, with differences between arteries and veins and between large and small arteries. Specific protein polarity and localization is essential for endothelial cells in responding to various extracellular stimuli. In this review, we examine protein localization in the endothelium of resistance arteries, with occasional references to other vessels for contrast, and how that polarization contributes to endothelial function and ultimately whole organism physiology. We highlight the protein localization on the luminal surface, discussing important physiological receptors and the glycocalyx. The protein polarization to the abluminal membrane is especially unique in small resistance arteries with the presence of the myoendothelial junction, a signaling microdomain that regulates vasodilation, feedback to smooth muscle cells, and ultimately total peripheral resistance. We also discuss the interendothelial junction, where tight junctions, adherens junctions, and gap junctions all convene and regulate endothelial function. Finally, we address planar cell polarity, or axial polarity, and how this is regulated by mechanosensory signals like blood flow.

mSphere of Influence: Compartmentalized cAMP signals in American trypanosomes.

Noelia Lander works on cell signaling in American trypanosomes and studies the role of cyclic adenosine monophosphate (cAMP) microdomains in environmental sensing and differentiation. In this mSphere of Influence, Dr. Lander reflects on three research articles in different eukaryotic models that had impacted on the way she thinks about the regulation of cAMP signals in Trypanosoma cruzi, the etiologic agent of Chagas disease. The articles "FRET biosensor uncovers cAMP nano-domains at ß-adrenergic targets that dictate precise tuning of cardiac contractility" (N. C. Surdo, M. Berrera, A. Koschinski, M. Brescia, et al., Nat Commun 8:15031, 2017, https://doi.org/10.1038/ncomms15031), "Cyclic AMP signaling and glucose metabolism mediate pH taxis by African trypanosomes" (S. Shaw, S. Knüsel, D. Abbühl, A. Naguleswaran, et al., Nat Commun 13:603, 2022, https://doi.org/10.1038/s41467-022-28293-w), and "Encystation stimuli sensing is mediated by adenylate cyclase AC2-dependent cAMP signaling in Giardia" (H. W. Shih, G. C. M. Alas, and A. R. Paredez, Nat Commun 14:7245, 2023, https://doi.org/10.1038/s41467-023-43028-1) influenced her current hypothesis that cAMP signals are generated in response to environmental cues leading to changes in membrane fluidity at the flagellar tip and the contractile vacuole complex of T. cruzi, structures where cAMP mediates key cellular processes for developmental progression.

Chimeric Antigen Receptor Signaling Domains Differentially Regulate Proliferation and Native T Cell Receptor Function in Virus-Specific T Cells.

The efficacy of T cells expressing chimeric antigen receptors (CARs) for solid tumors has been limited by insufficient CAR T cell expansion and persistence. The use of virus-specific T cells (VSTs) as carriers for CARs may overcome this limitation since CAR-VSTs can be boosted by viral vaccines or oncolytic viruses. However, there is limited understanding of the optimal combination of endodomains and their influence on the native T cell receptor (TCR) in VSTs. We therefore compared the function of GD2.CARs expressing the TCR zeta chain (ζ) alone or combined with endodomains from CD28 and 4-1BB in varicella zoster virus-specific (VZV) T cells. VZVSTs expressing GD2-CARs recognized VZV-derived peptides and killed GD2-expressing tumor cells. However, after repeated stimulation through their native TCR, the expansion of GD2-CAR.CD28ζ-VZVSTs was 3.3-fold greater (p < 0.001) than non-transduced VZVSTs, whereas GD2-CARζ- and GD2-CAR.41BBζ inhibited VZVST expansion (p < 0.01). Compared to control VZVSTs, GD2-CAR.ζ VZVSTs showed a greater frequency of apoptotic (p < 0.01) T cells, whereas prolonged downregulation of the native αß TCR was observed in GD2-CAR.41BBζ VZVSTs (p < 0.001). We confirmed that CD28ζ can best maintain TCR function by expressing GD2.CARs in Epstein-Barr virus-specific T cells and CD19-CARs in VZVSTs. In response to CAR stimulation VSTs with CD28ζ endodomains also showed the greatest expansion (6 fold > GD2-CAR.41BBζ VZVSTs (p < 0.001), however anti-tumor efficacy was superior in GD2-CAR.41BBζ-VZVSTs. These findings demonstrate that CAR signaling domains can enhance or diminish the function of the native TCR and indicate that only CD28ζ may preserve the function of the native TCR in tonically signaling CAR-VSTs.