RESUMO
The complex comprising serum response factor (SRF) and megakaryoblastic leukemia 1 protein (Mkl1) promotes myofibroblast differentiation during wound healing. SRF-Mkl1 is sensitive to the mechanical properties of the extracellular environment; but how cells sense and transduce mechanical cues to modulate SRF-Mkl1-dependent gene expression is not well understood. Here, we demonstrate that the nuclear lamina-associated inner nuclear membrane protein Emerin stimulates SRF-Mkl1-dependent gene activity in a substrate stiffness-dependent manner. Specifically, Emerin was required for Mkl1 nuclear accumulation and maximal SRF-Mkl1-dependent gene expression in response to serum stimulation of cells grown on stiff substrates but was dispensable on more compliant substrates. Focal adhesion area was also reduced in cells lacking Emerin, consistent with a role for Emerin in sensing substrate stiffness. Expression of a constitutively active form of Mkl1 bypassed the requirement for Emerin in SRF-Mkl1-dependent gene expression and reversed the focal adhesion defects evident in EmdKO fibroblasts. Together, these data indicate that Emerin, a conserved nuclear lamina protein, couples extracellular matrix mechanics and SRF-Mkl1-dependent transcription.
Assuntos
Proteínas de Membrana/genética , Proteínas Nucleares/genética , Fator de Resposta Sérica/genética , Transativadores/genética , Cicatrização/genética , Animais , Técnicas de Cultura de Células , Diferenciação Celular/genética , Adesões Focais/genética , Regulação da Expressão Gênica/genética , Humanos , Camundongos , Camundongos Knockout , Miofibroblastos/metabolismo , Membrana Nuclear/genética , Membrana Nuclear/metabolismo , Lâmina Nuclear/genética , Transdução de Sinais/genéticaRESUMO
Tumor-associated macrophages (TAM) play an important role in maintaining the immunosuppressive state of the tumor microenvironment (TME). High levels of CD163+ TAMs specifically are associated with poor prognosis in many solid tumor types. Targeting TAMs may represent a key approach in development of the next generation of cancer immune therapeutics. Members of the leukocyte immunoglobulin-like receptor B (LILRB) family, including LILRB2 (ILT4), are known to transmit inhibitory signals in macrophages and other myeloid cells. Leveraging bulk and single cell RNA-sequencing datasets, as well as extensive immunophenotyping of human tumors, we found that LILRB2 is highly expressed on CD163+ CD11b+ cells in the TME and that LILRB2 expression correlates with CD163 expression across many tumor types. To target LILRB2, we have developed JTX-8064, a highly potent and selective antagonistic mAb. JTX-8064 blocks LILRB2 binding to its cognate ligands, including classical and nonclassical MHC molecules. In vitro, JTX-8064 drives the polarization of human macrophages and dendritic cells toward an immunostimulatory phenotype. As a result, human macrophages treated with a LILRB2 blocker are reprogrammed to increase the activation of autologous T cells in co-culture systems. Furthermore, JTX-8064 significantly potentiates the activity of anti-PD-1 in allogeneic mixed lymphocyte reaction. In a human tumor explant culture, pharmacodynamic activity of JTX-8064 was observed in monotherapy and in combination with anti-PD-1. Collectively, our work provides strong translational and preclinical rationale to target LILRB2 in cancer.