Biology subjects

McInally, S. G.

Publications and source records attributed to McInally, S. G..

5 recordsLinked to original sources

bioRxiv2023

Length control emerges from cytoskeletal network geometry

Many cytoskeletal networks consist of individual filaments that are organized into elaborate higher order structures. While it is appreciated that the size and architecture of these networks are critical for their biological functions, much of the work investigating control over their assembly has focused on mechanisms that regulate the turnover of individual filaments through size-dependent feedback. Here, we propose a very different, feedback-independent mechanism to explain how yeast cells control the length of their actin cables. Our findings, supported by quantitative cell imaging and mathematical modeling, indicate that actin cable length control is an emergent property that arises from the cross-linked and bundled organization of the filaments within the cable. Using this model, we further dissect the mechanisms that allow cables to grow longer in larger cells, and propose that cell length-dependent tuning of formin activity allows cells to scale cable length with cell length. This mechanism is a significant departure from prior models of cytoskeletal filament length control and presents a new paradigm to consider how cells control the size, shape, and dynamics of higher order cytoskeletal structures. Significance StatementCells control the sizes of their cytoskeletal networks to ensure that these structures can efficiently perform their cellular functions. Until now, this ability has been attributed to molecular feedback mechanisms that control the rates at which individual filaments are assembled and disassembled. We find that size control of cytoskeletal networks does not require this type of feedback and can instead be encoded through the physical arrangement of the filaments within that network. These findings have important implications for understanding how the underlying geometry of higher order cytoskeletal networks contributes to cellular control over these structures.

cell biology↗

bioRxiv2023

Universal length fluctuations of actin structures found in cells

Actin is a key cytoskeletal protein that forms filaments that bundle into linear structures in vivo, which are involved in motility, signaling, and cell division. Despite the rapid turnover of individual actin monomers, these structures are often maintained at a specific length, which is important for their function. Length control is commonly attributed to length-dependent assembly or disassembly of the structure, whereby a stable length is achieved when the two opposing processes are balanced. Here we show that regardless of the nature of the length-dependent feedback, such "balance point" models predict a Gaussian distribution of lengths with a variance that is proportional to the steady state length. Contrary to this prediction, a reexamination of experimental measurements on the lengths of stereocilia, microvilli, actin cables, and filopodia reveals that the variance scales with the square of the steady state length. We propose a model in which the individual filaments in bundles undergo independent assembly dynamics, and the length of the bundle is set by the length of the longest filament. This model predicts a non-Gaussian distribution of bundle lengths with a variance that scales with the square of the steady state length. Our theory underscores the importance of crosslinking filaments into networks for size control of cytoskeleton structures.

cell biology↗

bioRxiv2021

Scaling of subcellular structures with cell length through decelerated growth

How cells tune the size of their subcellular parts to scale with cell size is a fundamental question in cell biology. Until now, most studies on the size control of organelles and other subcellular structures have focused on scaling relationships with cell volume, which can be explained by limiting pool mechanisms. Here, we uncover a distinct scaling relationship with cell length rather than volume, revealed by mathematical modeling and quantitative imaging of yeast actin cables. The extension rate of cables decelerates as they approach the rear of the cell, until cable length matches cell length. Further, the deceleration rate scales with cell length. These observations reveal a new mode of scaling that senses the linear dimensions of a cell. One Sentence SummaryAs actin cables in yeast cells grow longer, their extension rate decelerates, enabling the cable length to match cell length.

cell biology↗

bioRxiv2019

Length-dependent disassembly maintains four different flagellar lengths in Giardia

How flagellar length regulation is achieved in multiciliated eukaryotic cells with flagella of different equilibrium lengths is unknown. The protist Giardia lamblia is an ideal model to evaluate length regulation as it has flagella of four different lengths. Giardia axonemes have both non-membrane-bound and membrane-bound regions, but lack transition zones. Here we quantified the contributions of intraflagellar transport (IFT)-mediated assembly and kinesin-13-mediated disassembly to length control. IFT particles assemble and inject at Giardias flagellar pore complexes, which act as diffusion barriers functionally analogous to the transition zone to compartmentalize the membrane-bound regions of flagella. IFT-mediated assembly is length-independent as train size, speed, and injection frequencies are similar between flagella of different lengths. In Giardia, kinesin-13 mediates a length-dependent disassembly mechanism of length regulation to balance length-independent IFT-mediated assembly, resulting in different lengths. We anticipate that similar control mechanisms are widespread in multiciliated cells where cytoplasmic precursor pools are not limiting.

cell biology↗

bioRxiv2018

Robust and stable transcriptional repression in Giardia using CRISPRi

Giardia lamblia is a binucleate protistan parasite causing significant diarrheal disease worldwide. An inability to target Cas9 to both nuclei, combined with the lack of non-homologous end joining and markers for positive selection, has stalled the adaptation of CRISPR/Cas9-mediated genetic tools for this widespread parasite. CRISPR interference (CRISPRi) is a modification of the CRISPR/Cas9 system that directs catalytically inactive Cas9 (dCas9) to target loci for stable transcriptional repression. Using a Giardia nuclear localization signal to target dCas9 to both nuclei, we developed efficient and stable CRISPRi-mediated transcriptional repression of exogenous and endogenous genes in Giardia. Specifically, CRISPRi knockdown of kinesin-2a and kinesin-13 causes severe flagellar length defects that mirror defects with morpholino knockdown. Knockdown of the ventral disc MBP protein also causes severe structural defects that are highly prevalent and persist in the population more than five days longer than transient morpholino-based knockdown. By expressing two gRNAs in tandem to simultaneously knock down kinesin-13 and MBP, we created a stable dual knockdown strain with both flagellar length and disc defects. The efficiency and simplicity of CRISPRi in polyploid Giardia allows for rapid evaluation of knockdown phenotypes and highlights the utility of CRISPRi for emerging model systems.

microbiology↗