Toll-like receptor 4 (TLR4) is a significant mediator of cellular activation and antimicrobial responses through recognition of pathogen-derived and endogenous ligands. Its clinical relevance is underscored by associations with beneficial or deleterious outcomes in diverse disease contexts including infection, chronic inflammation, and cancer. The current paradigm holds that TLR4 signalling occurs sequentially, with cell surface-expressed TLR4 initially driving pro-inflammatory signalling. Subsequently, TLR4 endocytosis curbs this pro-inflammatory signalling and enables internalised TLR4 to signal from endosomes, resulting in type I Interferon (IFN) expression. Using murine primary macrophages and macrophage cell lines, we show that TLR4-induced type I IFN expression does not require TLR4 endocytosis, challenging the current viewpoint that these processes are inextricably linked. Moreover, we demonstrate that the presence and functionality of the TLR4 intracellular signalling domain are both indispensable for TLR4 endocytosis. This contrasts with current thinking that molecular regulators external to TLR4, such as CD14, control TLR4 endocytosis. Our data indicate that TLR4 endocytosis requires ubiquitin ligase activity, whereas components of canonical TLR signalling pathways are dispensable, implicating a thus far unrecognised mode of TLR4 signalling that governs activation-induced TLR4 endocytosis. Collectively, our data suggest that activation of cell surface-expressed TLR4 results in at least two distinct, TLR4-intrinsic signalling modes that independently control pro-inflammatory signalling and receptor endocytosis. Significantly, the latter is disconnected from endosomal TLR4 signalling. This revised understanding of TLR4 signalling as consisting of multiple functionally-distinct, non-sequential pathways may reveal opportunities for selective, disease context-specific amplification or restriction of TLR4 signalling pathways for beneficial therapeutic outcomes.