Cell size regulation arises from physical manifestations of cell proliferation and metabolic pathways. On one hand, coordination between these systems yields a constant cell size over generations to maintain cell size homeostasis. However, active regulation of cell size is crucial to physiology and to establish broad variation of cell sizes within an individual organism, and is accomplished via physical and biochemical pathways modulated by myriad intrinsic and extrinsic cues. In this review, we explore recent data elucidating the mechanobiological regulation of the volume of animal cells and its coordination with metabolic and proliferative pathways.

Septins are involved in many important cellular processes, and septin dysfunction has been implicated in various pathologies, such as cancer. Like other components of the cytoskeleton -F-actin, microtubules, and intermediate filaments-septins can self-assemble into filaments and higher-order structures. These non-polar filaments are assembled from complex and variable multimeric building blocks. Septins exhibit a distinct preference for interacting with actin and microtubule structures, particularly at the interface with cellular membrane. Although they are crucial for many vital cellular functions and are frequently observed at prominent cellular structures like stress fibers, cilia, and neuronal processes, our understanding of the regulation of septin filament dynamics and the organized assembly of higher-order structures remains limited. However, recent insights into the architecture of septin filaments, the structure of crucial septin domains, and their interactions with other cellular components (F-actin, microtubules, membranes) and regulatory proteins may now pave the way for rapid progress.

Cell biology emerges from spatiotemporally coordinated molecular processes. Recent advances in live-cell microscopy, fueled by a surge in optical, molecular, and computational technologies, have enabled dynamic observations from single molecules to whole organisms. Despite technological leaps, there is still an untapped opportunity to fully leverage their capabilities toward biological insight. We highlight how single-molecule imaging has transformed our understanding of biological processes, with a focus on chromatin organization and transcription in the nucleus. We describe how this was enabled by the close integration of new imaging techniques with analysis tools and discuss the challenges to make a comparable impact at larger scales from organelles to organisms. By highlighting recent successful examples, we describe an outlook of ever-increasing data and the need for seamless integration between dataset visualization and quantification to realize the full potential warranted by advances in new imaging technologies.

Physical parameters such as tissue interplay forces, luminal pressure, fluid flow, temperature, and electric fields are crucial regulators of embryonic morphogenesis. While significant attention has been given to cellular and molecular responses to these physical parameters, their roles in morphogenesis are not yet fully elucidated. This is largely due to a shortage of methods for spatiotemporal modulation and direct quantitative perturbation of physical parameters in embryos. Recent advancements addressing these challenges include microscopes equipped with devices to apply and adjust forces, direct perturbation of luminal pressure, and the application of micro-forces to targeted cells and cilia in vivo. These methods are critical for unveiling morphogenesis mechanisms, highlighting the importance of integrating molecular and physical approaches for a comprehensive understanding of morphogenesis.

Since the advent of Hi-C in 2009, a plethora of high-throughput sequencing methods have emerged to profile the three-dimensional (3D) organization of eukaryotic genomes, igniting the era of 3D genomics. In recent years, the genomic resolution achievable by these approaches has dramatically increased and several single-cell versions of Hi-C have been developed. Moreover, a new repertoire of tools not based on proximity ligation of digested chromatin has emerged, enabling the investigation of the higher-order organization of chromatin in the nucleus. In this review, we summarize the expanding portfolio of 3D genomic technologies, highlighting recent developments and applications from the past three years. Lastly, we present an outlook of where this technology-driven field might be headed.

Many solid tumors exhibit significant genetic, cellular, and biophysical heterogeneity which dynamically evolves during disease progression and after treatment. This constant flux in cell composition, phenotype, spatial relationships, and tissue properties poses significant challenges in accurately diagnosing and treating patients. Much of the complexity lies in unraveling the molecular changes in different tumor compartments, how they influence one another in space and time and where vulnerabilities exist that might be appropriate to target therapeutically. Recent advances in spatial profiling tools and technologies are enabling new insight into the underlying biology of complex tumors, creating a greater understanding of the intricate relationship between cell types, states, and the microenvironment. Here we reflect on some recent discoveries in this area, where the key knowledge and technology gaps lie, and the advancements in spatial measurements and in vitro models for the study of spatial intratumoral heterogeneity.

The dynamic actin cytoskeleton contributes to many critical biological processes by providing the structural support underlying the morphology of most cells, facilitating intracellular transport, and generating forces required for cell motility and division. To execute many of these functions, actin monomers polymerize into polarized filaments that display different structural and biochemical properties at each end. Filament dynamics are regulated by diverse regulatory proteins which collaborate to dictate rates of elongation and disassembly, particularly at the fast-growing barbed (plus) end. This review highlights the biochemical mechanisms of six barbed end regulatory proteins: formin, profilin, capping protein, IQGAP1, cyclase-associated protein, and twinfilin. We discuss how individual proteins influence actin dynamics and how several intriguing complex assemblies influence the polymerization fate of actin filaments. Understanding these mechanisms offers insights into how actin is regulated in essential cell processes and dysregulated in disease.

Legume roots allow intracellular infections of rhizobia to establish the mutualistic root nodule symbiosis. During this colonization event, specialized and membrane-defined infection threads provide the host-controlled path for the bacteria through the multilayered root tissue to reach a newly developing organ, the root nodule. On this way, bacteria have to propagate transcellularly and thus overcome cell wall barriers. This process not only requires continuous molecular surveillance of the invading microbe but also structural adaptations of the extracellular matrix components in a spatially confined manner leading to the formation of a novel compartment that we term the "transcellular passage cleft" (TPC). Here, we review the molecular mechanisms and signaling events around the TPC and propose a step-wise model for TPC formation.

As animals develop, molecules, cells, and cell ensembles move in beautifully orchestrated choreographies. Movement at each of these scales requires generation of mechanical force. In eukaryotic cells, the actomyosin cytoskeleton generates mechanical forces. Continuous advances in in vivo microscopy have enabled visualization and quantitative assessment of actomyosin dynamics and force generation, within and across cells, in living embryos. Recent studies reveal that actomyosin networks can form periodic waves in vivo. Here, we highlight contributions of actomyosin waves to molecular transport, cell movement, and cell coordination in developing embryos.

S.J. Gould and R. Lewontin in their famous "Spandrels paper" (1979) argued that many anatomical elements arise in evolution not due to their "current utility" but rather due to other "reasons for origin", such as other developmental processes, physical constraints and mechanical forces. Here, in the same spirit, we argue that a variety of molecular processes, physical constraints, and mechanical forces, alone or together, generate structures that are detectable in the cell nucleus, yet these structures themselves may not carry any specific function, being a mere reflection of processes that produced them.

In recent years, the role of 53BP1 as a cell cycle regulator has come into the spotlight. 53BP1 is best understood for its role in controlling DNA double-strand break repair. However, 53BP1 was initially discovered as an interaction partner of the tumor suppressor p53, which proved to be independent of DNA repair. The importance of this interaction is becoming increasingly clear. 53BP1 responds to mitotic stress, which prolongs mitosis, or to DNA damage and triggers the stabilization of p53 by the deubiquitinase USP28 to stop the proliferation of potentially damaged cells. The ability of 53BP1 to respond to mitotic stress or DNA damage is controlled by cell cycle-specific post-translational modifications and is therefore restricted to specific cell cycle phases. 53BP1-mediated p53 activation is likely involved in tumor suppression and is associated with genetic diseases such as primary microcephaly. This review emphasizes the importance of these mechanisms for the development and maintenance of healthy tissues.

In actively dividing eukaryotic cells, the nuclear envelope membrane (NEM) expands during the cell cycle to accommodate increases in nuclear volume and formation of two nuclei as a cell passes through mitosis to form daughter cells. NEM expansion is driven by glycerophospholipid (GPL) synthesis that is regulated by the lipin family of phosphatidic acid phosphatases (PAPs). How, and when during the cell cycle, PAPs regulate membrane expansion differs between organisms undergoing a closed or open mitosis. Here, we discuss recent studies that shed light on the mechanisms of NE expansion. Moreover, we examine evidence that NEM expansion not only employs GPLs synthesized in the ER but also lipids whose synthesis is regulated by events at the inner nuclear membrane.

Nucleocytoplasmic transport is a basic cellular reaction that plays an important role in regulating cell physiology in eukaryotic cells. Here we show that the identification of one nucleocytoplasmic transport pathway led to the notification of intracellular reaction that has not been acknowledged. Hikeshi was originally identified as a nuclear import carrier of heat stress-induced nuclear import of molecular chaperone Hsp70. We now know that Hikeshi mediates nuclear import of Hsp70 at a variety of different cellular conditions, such as at normal conditions, at proteotoxic conditions, during differentiation, and probably more. Recent studies gradually revealed the physiological significances of Hikeshi-mediated nuclear import of Hsp70.

TERRA long noncoding RNAs play key roles in telomere function and maintenance. They can orchestrate telomeric chromatin remodeling, regulate telomere maintenance by telomerase and homology-directed repair, and they participate in the telomeric DNA damage response. TERRA associates with chromosome ends through base-pairing forming R-loops, which are mediated by the RAD51 DNA recombinase and its partner RAD51AP1. Telomeric R-loops interfere with replication fork progression, stimulating a switch of telomere maintenance from semiconservative DNA replication to homology-directed repair (HDR). The latter mechanism is exploited by a subset of cancer cells that lack telomerase, referred to as ALT. In addition, TERRA stimulates HDR at short telomeres during aging, delaying cellular senescence. During carcinogenesis, when cells with eroded telomeres enter replicative crisis, TERRA acts as a signaling molecule to mediate autophagic cell death.

The efficient distribution of oxygen and metabolites is critical for embryonic development and growth as well as tissue homeostasis. This is achieved by endothelial cells forming and maintaining a closed, circulatory network of tubular blood vessels. Endothelial cells are highly plastic cells with the capability to generate diverse dynamic responses at different stages of vessel development in order to build vessel networks of tissue-specific patterns and morphologies. In this review, we discuss new conceptual advances gained from in vitro and in vivo models of angiogenesis on the control of endothelial cell dynamics. We highlight the complex interplay between mechanical cues, actin cytoskeleton and endothelial behaviors, and the emerging importance of hydrostatic pressure in complementing actin-dependent mechanisms to regulate endothelial cell mechanics and angiogenesis. Understanding these processes provides insights into vascular repair and regeneration mechanisms.

Epithelial cells adhere to each other via intercellular junctions that can be classified into bicellular junctions and tricellular contacts (vertices). Epithelial morphogenesis involves cell rearrangement and requires remodeling of bicellular junctions and vertices. Although our understanding of how bicellular junction mechanics drive epithelial morphogenesis has advanced, the mechanisms underlying vertex remodeling during this process have only received attention recently. In this review, we outline recent progress in our understanding of how cells reorganize cell adhesion and the cytoskeleton to trigger the displacement and resolution of cell vertices. We will also discuss how cells achieve the optimal balance between the structural flexibility and stability of their vertices. Finally, we introduce new modeling frameworks designed to analyze mechanics at cell vertices. Integration of live imaging and modeling techniques is providing new insights into the active roles of cell vertices during epithelial morphogenesis.

The Arf family GTPases are regulators of eukaryotic cellular organization, functioning in the secretory and endocytic pathways, in cilia and flagella, in cytoskeleton dynamics, and in lipid metabolism. We describe the evolution of this protein family and its well-studied regulators. The last eukaryotic common ancestor had fifteen members, and the current complement of Arf GTPases has been sculpted by gene loss and gene duplications since that point. Some Arf family GTPases (such as those that recruit vesicle coats in the secretory pathway) are present in virtually all eukaryotes, whereas others (such as those functioning in cilia/flagella) have a more limited distribution. A challenge for the future is understanding the full spectrum of Arf family functions throughout eukaryotes.

Stem cell models for early mammalian development offer new experimental opportunities to access spatio-temporal details of the cell-cell interactions that govern cell differentiation and tissue patterning. This review summarizes recent studies that have used stem cell models to investigate the spatial range of developmental cell-cell communication systems. A key message from these works is that important biochemical signals for cell differentiation in these systems, such as Nodal and fibroblast growth factors (FGFs), often act over short distances of only a few cell diameters. The formation of long-range patterns at the tissue scale associated with these signals then results from signal relays and cell rearrangements. The modular view of differentiation and patterning emerging from research on stem cell models can offer a fresh perspective on the corresponding processes in the embryo.

Research in the areas of organelle dynamics, cytoskeletal interactions, membrane protrusions, and cell motility relies heavily on live-cell imaging. These structures continuously move about in complex patterns and imaging them live at sufficient temporal resolutions as well as for durations long enough to extract significant number of events is an absolute necessity. Capturing most of the sub-cellular dynamics in whole cell volumes was beyond reach due to the lack of balance between reduced photo-toxicity, time resolution, and the required spatial resolution in dominant imaging modalities like point scanning confocal and spinning disc confocal microscopy. In the last few years, a plethora of light-sheet geometries have emerged, pushing the limits of measurements. In this review, we will focus on a subset of light-sheet modalities that are most suited to studying live, sub-cellular dynamics in whole-cell volumes.