Allogeneic hematopoietic stem cell transplantation (alloSCT) is an important curative therapy for high-risk hematological malignancies, but the development of severe and/or steroid-refractory acute graft-versus-host disease (aGVHD) remains a significant limitation to optimal outcomes. New approaches to prevent and treat aGVHD remain an unmet need that can be best addressed by understanding the complex disease pathophysiology. It is now clear that chemoradiotherapy used prior to alloSCT induces the release of endogenous alarmins (eg, HMGB-1, ATP, IL-1α, IL-33) from recipient tissue. Exogenous pathogen-derived molecules (eg, lipopolysaccharide, nucleic acids) also translocate from the gastrointestinal tract lumen. Together, these danger signals activate antigen-presenting cells (APCs) to efficiently present alloantigen to donor T cells while releasing cytokines (eg, interleukin-12 [IL-12], IL-23, IL-6, IL-27, IL-10, transforming growth factor-β) that expand and differentiate both pathogenic and regulatory donor T cells. Concurrent costimulatory signals at the APC–T-cell interface (eg, CD80/CD86-CD28, CD40-CD40L, OX40L-OX40, CD155/CD112-DNAM-1) and subsequent coinhibitory signals (eg, CD80/CD86-CTLA4, PDL1/2-PD1, CD155/CD112-TIGIT) are critical to the acquisition of effector T-cell function and ensuing secretion of pathogenic cytokines (eg, IL-17, interferon-γ, tissue necrosis factor, granulocyte-macrophage colony-stimulating factor) and cytolytic degranulation pathway effectors (eg, perforin/granzyme). This review focuses on the combination of cytokine and costimulatory networks at the T-cell surface that culminates in effector function and subsequent aGVHD in target tissue. Together, these pathways now represent robust and clinically tractable targets for preventing the initiation of deleterious immunity after alloSCT.