Colony context and size-dependent compensation mechanisms give rise to variations in nuclear growth trajectories.

To investigate the fundamental question of how cellular variations arise across spatiotemporal scales in a population of identical healthy cells, we focused on nuclear growth in hiPS cell colonies as a model system. We generated a 3D timelapse dataset of thousands of nuclei over multiple days, and developed open-source tools for image and data analysis and an interactive timelapse viewer for exploring quantitative features of nuclear size and shape. We performed a data-driven analysis of nuclear growth variations across timescales. We found that individual nuclear volume growth trajectories arise from short timescale variations attributable to their spatiotemporal context within the colony. We identified a strikingly time-invariant volume compensation relationship between nuclear growth duration and starting volume across the population. Notably, we discovered that inheritance plays a crucial role in determining these two key nuclear growth features while other growth features are determined by their spatiotemporal context and are not inherited.

Specialized cells that sense tissue mechanics to regulate Drosophila morphogenesis.

Shaping of developing organs requires dynamic regulation of force and resistance to achieve precise outcomes, but how organs monitor tissue mechanical properties is poorly understood. We show that in developing Drosophila follicles (egg chambers), a single pair of cells performs such monitoring to drive organ shaping. These anterior polar cells secrete a matrix metalloproteinase (MMP) that specifies the appropriate degree of tissue elongation, rather than hyper- or hypo-elongated organs. MMP production is negatively regulated by basement membrane (BM) mechanical properties, which are sensed through focal adhesion signaling and autonomous contractile activity; MMP then reciprocally regulates BM remodeling, particularly at the anterior region. Changing BM properties at remote locations alone is sufficient to induce a remodeling response in polar cells. We propose that this small group of cells senses both local and distant stiffness cues to produce factors that pattern the organ's BM mechanics, ensuring proper tissue shape and reproductive success.

Surface Area to Volume Ratio: A Natural Variable for Bacterial Morphogenesis.

An immediately observable feature of bacteria is that cell size and shape are remarkably constant and characteristic for a given species in a particular condition, but vary quantitatively with physiological parameters such as growth rate, indicating both genetic and environmental regulation. However, despite decades of research, the molecular mechanisms underlying bacterial morphogenesis have remained incompletely characterized. We recently demonstrated that a wide range of bacterial species exhibit a robust surface area to volume ratio (SA/V) homeostasis. Because cell size, shape, and SA/V are mathematically interconnected, if SA/V is indeed the natural variable that cells actively monitor, this finding has critical implications for our understanding of bacterial morphogenesis, placing fundamental constraints on the sizes and shapes that cells can adopt. In this Opinion article we discuss the broad implications that this novel perspective has for the field of bacterial growth and morphogenesis.

Relative Rates of Surface and Volume Synthesis Set Bacterial Cell Size.

Many studies have focused on the mechanisms underlying length and width determination in rod-shaped bacteria. Here, we focus instead on cell surface area to volume ratio (SA/V) and demonstrate that SA/V homeostasis underlies size determination. We propose a model whereby the instantaneous rates of surface and volume synthesis both scale with volume. This model predicts that these relative rates dictate SA/V and that cells approach a new steady-state SA/V exponentially, with a decay constant equal to the volume growth rate. To test this, we exposed diverse bacterial species to sublethal concentrations of a cell wall biosynthesis inhibitor and observed dose-dependent decreases in SA/V. Furthermore, this decrease was exponential and had the expected decay constant. The model also quantitatively describes SA/V alterations induced by other chemical, nutritional, and genetic perturbations. We additionally present evidence for a surface material accumulation threshold underlying division, sensitizing cell length to changes in SA/V requirements.

A Caulobacter†MreB mutant with irregular cell shape exhibits compensatory widening to maintain a preferred surface area to volume ratio.

Rod-shaped bacteria typically elongate at a uniform width. To investigate the genetic and physiological determinants involved in this process, we studied a mutation in the morphogenetic protein MreB in Caulobacter crescentus that gives rise to cells with a variable-width phenotype, where cells have regions that are both thinner and wider than wild-type. During growth, individual cells develop a balance of wide and thin regions, and mutant MreB dynamically localizes to poles and thin regions. Surprisingly, the surface area to volume ratio of these irregularly shaped cells is, on average, very similar to wild-type. We propose that, while mutant MreB localizes to thin regions and promotes rod-like growth there, wide regions develop as a compensatory mechanism, allowing cells to maintain a wild-type-like surface area to volume ratio. To support this model, we have shown that cell widening is abrogated in growth conditions that promote higher surface area to volume ratios, and we have observed individual cells with high ratios return to wild-type levels over several hours by developing wide regions, suggesting that compensation can take place at the level of individual cells.

A general assay for monitoring the activities of protein tyrosine phosphatases in living eukaryotic cells.

Protein tyrosine phosphatases (PTPs) are key signal-transduction regulators and have emerged as potential drug targets for inhibitor design. Here we report a yeast-based assay that provides a general means of assessing the activity and/or inhibition of essentially any classical PTP in living cells. The assay uses the activity of an exogenously expressed PTP to counter the activity of a coexpressed and toxic tyrosine kinase, such that only active PTPs are capable of rescuing growth. PTP activity gives rise to both increased growth and decreased phosphotyrosine levels; cellular PTP activity can therefore be monitored by either yeast-growth curves or anti-phosphotyrosine Western blots. We show that four PTPs (TCPTP, Shp2, PEST, PTPα) are capable of rescuing the effects of v-Src toxicity. Since these PTPs are chosen from four distinct subfamilies, it is likely that biologically and medicinally important PTPs from other subfamilies can similarly function in the cellular PTP assay. Because many small-molecule PTP inhibitors fail to penetrate cell membranes effectively, this cell-based assay has the potential to serve as a useful screening tool for determining the cellular efficacy of candidate inhibitors in a more biologically relevant context than can be provided by an in vitro PTP assay.

The effect of chromosome geometry on genetic diversity.

Although organisms with linear chromosomes must solve the problem of fully replicating their chromosome ends, this chromosome configuration has emerged repeatedly during bacterial evolution and is evident in three divergent bacterial phyla. The benefit usually ascribed to this topology is the ability to boost genetic variation through increased recombination. But because numerous processes can impact linkage disequilibrium, such an effect is difficult to assess by comparing across bacterial taxa that possess different chromosome topologies. To test directly the contribution of chromosome architecture to genetic diversity and recombination, we examined sequence variation in strains of Agrobacterium Biovar 1, which are unique among sequenced bacteria in having both a circular and a linear chromosome. Whereas the allelic diversity among strains is generated principally by mutations, intragenic recombination is higher within genes situated on the circular chromosome. In contrast, recombination between genes is, on average, higher on the linear chromosome, but it occurs at the same rate as that observed between genes mapping to the distal portion of the circular chromosome. Collectively, our findings indicate that chromosome topology does not contribute significantly to either allelic or genotypic diversity and that the evolution of linear chromosomes is not based on a facility to recombine.