Integrative modeling enables structure determination for large macromolecular assemblies by combining data from multiple experiments with theoretical and computational predictions. Recent advancements in AI-based structure prediction and cryo electron-microscopy have sparked renewed enthusiasm for integrative modeling; structures from AI-based methods can be integrated with in situ maps to characterize large assemblies. This approach previously allowed us and others to determine the architectures of diverse macromolecular assemblies, such as nuclear pore complexes, chromatin remodelers, and cell–cell junctions. Experimental data spanning several scales was used in these studies, ranging from high-resolution data, such as X-ray crystallography and AlphaFold structure, to low-resolution data, such as cryo-electron tomography maps and data from co-immunoprecipitation experiments. Two recurrent modeling challenges emerged across a range of studies. First, these assemblies contained significant fractions of disordered regions, necessitating the development of new methods for modeling disordered regions in the context of ordered regions. Second, methods needed to be developed to utilize the information from cryo-electron tomography, a timely challenge as structural biology is increasingly moving towards in situ characterization. Here, we recapitulate recent developments in the modeling of disordered proteins and the analysis of cryo-electron tomography data and highlight other opportunities for method development in the context of integrative modeling.

In situ structural biology with cryo-electron tomography (cryo-ET) and subtomogram averaging (StA) is evolving as a major method to understand the structure, function, and interactions of biological molecules in cells in a single experiment. Since its inception, the method has matured with some stellar highlights and with further opportunities to broaden its applications. In this short review, I want to provide a personal perspective on the developments in cryo-ET as I have seen it for the last ~20 years and outline the major steps that led to its success. This perspective highlights cryo-ET with my eyes as a junior researcher and my view on the present and past developments in hardware and software for in situ structural biology with cryo-ET.

Host-virus interactions are critically important for various stages of the viral replication cycle. The reliance of viruses on the host factors for their entry, replication, and maturation processes can be exploited for the development of antiviral therapeutics. Thus, the identification and characterization of such viral-host dependency factors has been an attractive area of research to provide novel antiviral targets. Traditional proteomic efforts based on affinity purification of protein complexes from cell lysates are limited to detecting strong and stable interactions. In this perspective, we discuss the integration of two latest proteomic techniques, based on in situ proximity labelling and chemical crosslinking methods, to uncover host-virus protein–protein interactions in living cells.