Background and Aims: Immune-checkpoint inhibitors target the membrane protein; however, the role of membrane proteins in antitumor immunity remains poorly elucidated. In this study, we aimed to explore the role of membrane proteins and unearth potential membrane proteins that can be targeted. Approach and Results: We initially screened prognosis-related membrane proteins based on The Cancer Genome Atlas Program and International Cancer Genome Consortium databases. Whole-gene, T-cell–specific or CD8T-cell–specific integral membrane protein 2A (ITM2A) knockout mice were constructed and used for orthotopic transplantation or as plasmid-derived spontaneous hepatocellular carcinoma (HCC) models to explore the role of ITM2A in the tumor microenvironment (TME). Through single-cell RNA sequencing (scRNA-seq), TimiGP analysis, molecular dynamics simulations, and biochemical experiments, the molecular mechanisms underlying the MSR1–ITM2A–TCR signaling regulatory axis were explored. Finally, bioengineering technologies were used to design and construct a new CD4T-targeted antibody–drug conjugate (ADC).High ITM2A expression in HCC indicated a superior prognosis, which was associated with richer immune cell infiltration in the TME. The results of the scRNA-seq of the HCC model in genetically knocked-out mice suggest that ITM2A influences the TCR signaling of tumor-infiltrating lymphocytes in the TME. This inference was confirmed in conditional ITM2A knockout mice. MSR1 ⁺ macrophages induced CD4T exhaustion by disrupting the ITM2A–ZAP70 axis during TCR activation. ADC (ZEA-αCD4) treatment effectively inhibited tumor growth, independent of the classical αPDL1 immunotherapy. Conclusions: MSR1 ⁺ macrophages promote tumor-infiltrating ITM2A ⁺ CD4T exhaustion differentiation by regulating the ITM2A–ZAP70 axis. ZEA-αCD4 specifically targeted the MSR1–ITM2A interaction to activate antitumor immunity and improve the efficacy of αPDL1 immunotherapy.

The m⁶A RNA methylation pathway plays a critical role in host antiviral defense. Host cells employ m⁶A readers such as YTHDF2 to regulate viral RNA fate through diverse mechanisms, including degradation, translational control, and immune recognition. However, we found that YTHDF2 is essential for SARS-CoV-2 replication, suggesting that a virus may exploit this host machinery to its advantage. Through integrative RNA-proteome analysis, we identified the SARS-CoV-2 nucleocapsid (N) transcript as the most heavily m⁶A-modified viral transcript and a direct interactor of YTHDF2. The N protein forms a complex with YTHDF2 in the cytoplasm and redirects this host RNA decay machinery toward host antiviral transcripts. N suppresses ISG15, IFIT1, MX1 and pro-inflammatory cytokines in a largely YTHDF2-dependent manner, an effect that is lost in YTHDF2-knockout cells. These findings reveal a viral immune evasion strategy wherein a viral protein actively hijacks an m⁶A reader to silence antiviral gene expression, establishing the N-YTHDF2 axis as a therapeutic target against SARS-CoV-2 and other coronaviruses.

Branching morphogenesis shapes the epithelial tissue architecture as proliferative branches navigate the extracellular matrix (ECM) microenvironment. In the mammary gland, pubertal terminal end buds (TEBs) drive stochastic ductal invasion to build a tree-like network. Although stem cell programs and hormonal regulation have been extensively characterized, the geometric principles underlying branch adaptation to ECM mechanics during morphogenesis remain incompletely defined. Here, using in vivo whole-mount imaging and ex vivo 3D organoid live imaging, we show that the mammary bifurcation angle occupies a characteristic range and, in organoids, that it oscillates dynamically during branch morphogenesis. We show that branch angles maintain a confined dynamic range with regular oscillations during both bud bifurcation and branch elongation, thereby supporting efficient branching morphogenesis. These angle dynamics are independent of major mammary-specific developmental growth and hormonal signaling, pointing to non-endocrine microenvironmental cues as potential regulators. Integrative analysis of branched epithelial RNA-seq and in vivo TEB microarray data further reveals the remodeling of cell adhesion pathways and epithelial–ECM interactions during network formation, with matrix metalloproteinase 14 (MMP14) specifically enriched in branched epithelia as a promising candidate mediator. Pharmacological inhibition of MMP14 disrupted branch angle oscillations and impaired organoid branching growth. Taken together, our findings, derived from murine tissue and organoid models, identify the homeostatic branch angle as a previously unrecognized biophysical signature of mammary branching that is maintained, in organoids, through MMP14-mediated ECM remodeling.