Myosin XI, a model of its conserved role in plant cell tip growth.

In eukaryotic cells, organelle and vesicle transport, positioning, and interactions play crucial roles in cytoplasmic organization and function. These processes are governed by intracellular trafficking mechanisms. At the core of that trafficking, the cytoskeleton and directional transport by motor proteins stand out as its key regulators. Plant cell tip growth is a well-studied example of cytoplasm organization by polarization. This polarization, essential for the cell's function, is driven by the cytoskeleton and its associated motors. This review will focus on myosin XI, a molecular motor critical for vesicle trafficking and polarized plant cell growth. We will center our discussion on recent data from the moss Physcomitrium patens and the liverwort Marchantia polymorpha. The biochemical properties and structure of myosin XI in various plant species are discussed, highlighting functional conservation across species. We further explore this conservation of myosin XI function in the process of vesicle transport in tip-growing cells. Existing evidence indicates that myosin XI actively organizes actin filaments in tip-growing cells by a mechanism based on vesicle clustering at their tips. A hypothetical model is presented to explain the essential function of myosin XI in polarized plant cell growth based on vesicle clustering at the tip. The review also provides insight into the in vivo localization and dynamics of myosin XI, emphasizing its role in cytosolic calcium regulation, which influences the polymerization of F-actin. Lastly, we touch upon the need for additional research to elucidate the regulation of myosin function.

Measurement of the persistence length of cytoskeletal filaments using curvature distributions.

Cytoskeletal filaments, such as microtubules and actin filaments, play important roles in the mechanical integrity of cells and the ability of cells to respond to their environment. Measuring the mechanical properties of cytoskeletal structures is crucial for gaining insight into intracellular mechanical stresses and their role in regulating cellular processes. One of the ways to characterize these mechanical properties is by measuring their persistence length, the average length over which filaments stay straight. There are several approaches in the literature for measuring filament deformations, such as Fourier analysis of images obtained using fluorescence microscopy. Here, we show how curvature distributions can be used as an alternative tool to quantify biofilament deformations, and investigate how the apparent stiffness of filaments depends on the resolution and noise of the imaging system. We present analytical calculations of the scaling curvature distributions as a function of filament discretization, and test our predictions by comparing Monte Carlo simulations with results from existing techniques. We also apply our approach to microtubules and actin filaments obtained from in vitro gliding assay experiments with high densities of nonfunctional motors, and calculate the persistence length of these filaments. The presented curvature analysis is significantly more accurate compared with existing approaches for small data sets, and can be readily applied to both in vitro and in vivo filament data through the use of the open-source codes we provide.

In vivo interactions between myosin XI, vesicles and filamentous actin are fast and transient in Physcomitrella patens.

The actin cytoskeleton and active membrane trafficking machinery are essential for polarized cell growth. To understand the interactions between myosin XI, vesicles and actin filaments in vivo, we performed fluorescence recovery after photobleaching and showed that the dynamics of myosin XIa at the tip of the spreading earthmoss Physcomitrella patens caulonemal cells are actin-dependent and that 50% of myosin XI is bound to vesicles. To obtain single-particle information, we used variable-angle epifluorescence microscopy in protoplasts to demonstrate that protein myosin XIa and VAMP72-labeled vesicles localize in time and space over periods lasting only a few seconds. By tracking data with Hidden Markov modeling, we showed that myosin XIa and VAMP72-labeled vesicles exhibit short runs of actin-dependent directed transport. We also found that the interaction of myosin XI with vesicles is short-lived. Together, this vesicle-bound fraction, fast off-rate and short average distance traveled seem be crucial for the dynamic oscillations observed at the tip, and might be vital for regulation and recycling of the exocytosis machinery, while simultaneously promoting vesicle focusing and vesicle secretion at the tip, necessary for cell wall expansion.

Myosin XI drives polarized growth by vesicle focusing and local enrichment of F-actin in Physcomitrium patens.

In tip-growing plant cells, growth results from myosin XI and F-actin-mediated deposition of cell wall polysaccharides contained in secretory vesicles. Previous evidence showed that myosin XI anticipates F-actin accumulation at the cell's tip, suggesting a mechanism where vesicle clustering via myosin XI increases F-actin polymerization. To evaluate this model, we used a conditional loss-of-function strategy by generating moss (Physcomitrium patens) plants harboring a myosin XI temperature-sensitive allele. We found that loss of myosin XI function alters tip cell morphology, vacuolar homeostasis, and cell viability but not following F-actin depolymerization. Importantly, our conditional loss-of-function analysis shows that myosin XI focuses and directs vesicles at the tip of the cell, which induces formin-dependent F-actin polymerization, increasing F-actin's local concentration. Our findings support the role of myosin XI in vesicle focusing, possibly via clustering and F-actin organization, necessary for tip growth, and deepen our understanding of additional myosin XI functions.

Quantitative cell biology of tip growth in moss.

KEY MESSAGE: Here we review, from a quantitative point of view, the cell biology of protonemal tip growth in the model moss Physcomitrium patens. We focus on the role of the cytoskeleton, vesicle trafficking, and cell wall mechanics, including reviewing some of the existing mathematical models of tip growth. We provide a primer for existing cell biological tools that can be applied to the future study of tip growth in moss. Polarized cell growth is a ubiquitous process throughout the plant kingdom in which the cell elongates in a self-similar manner. This process is important for nutrient uptake by root hairs, fertilization by pollen, and gametophyte development by the protonemata of bryophytes and ferns. In this review, we will focus on the tip growth of moss cells, emphasizing the role of cytoskeletal organization, cytoplasmic zonation, vesicle trafficking, cell wall composition, and dynamics. We compare some of the existing knowledge on tip growth in protonemata against what is known in pollen tubes and root hairs, which are better-studied tip growing cells. To fully understand how plant cells grow requires that we deepen our knowledge in a variety of forms of plant cell growth. We focus this review on the model plant Physcomitrium patens, which uses tip growth as the dominant form of growth at its protonemal stage. Because mosses and vascular plants shared a common ancestor more than 450 million years ago, we anticipate that both similarities and differences between tip growing plant cells will provide mechanistic information of tip growth as well as of plant cell growth in general. Towards this mechanistic understanding, we will also review some of the existing mathematical models of plant tip growth and their applicability to investigate protonemal morphogenesis. We attempt to integrate the conclusions and data across cell biology and physical modeling to our current state of knowledge of polarized cell growth in P. patens and highlight future directions in the field.

Rab-E and its interaction with myosin XI are essential for polarised cell growth.

The fundamental process of polarised exocytosis requires the interconnected activity of molecular motors trafficking vesicular cargo within a dynamic cytoskeletal network. In plants, few mechanistic details are known about how molecular motors, such as myosin XI, associate with their secretory cargo to support the ubiquitous processes of polarised growth and cell division. Live-cell imaging coupled with targeted gene knockouts and a high-throughput RNAi assay enabled the first characterisation of the loss of Rab-E function. Yeast two-hybrid and subsequent in silico structural prediction uncovered a specific interaction between Rab-E and myosin XI that is conserved between P. patens and A. thaliana. Rab-E co-localises with myosin XI at sites of active exocytosis, and at the growing tip both proteins are spatiotemporally coupled. Rab-E is required for normal plant growth in P. patens and the rab-E and myosin XI phenotypes are rescued by A. thaliana's Rab-E1c and myosin XI-K/E, respectively. Both PpMyoXI and AtMyoXI-K interact with PpRabE14, and the interaction is specifically mediated by PpMyoXI residue V1422. This interaction is required for polarised growth. Our results suggest that the interaction of Rab-E and myosin XI is a conserved feature of polarised growth in plants.

Robust Survival-Based RNA Interference of Gene Families Using in Tandem Silencing of Adenine Phosphoribosyltransferase.

RNA interference (RNAi) enables flexible and dynamic interrogation of entire gene families or essential genes without the need for exogenous proteins, unlike CRISPR-Cas technology. Unfortunately, isolation of plants undergoing potent gene silencing requires laborious design, visual screening, and physical separation for downstream characterization. Here, we developed an adenine phosphoribosyltransferase (APT)-based RNAi technology (APTi) in Physcomitrella patens that improves upon the multiple limitations of current RNAi techniques. APTi exploits the prosurvival output of transiently silencing APT in the presence of 2-fluoroadenine, thereby establishing survival itself as a reporter of RNAi. To maximize the silencing efficacy of gene targets, we created vectors that facilitate insertion of any gene target sequence in tandem with the APT silencing motif. We tested the efficacy of APTi with two gene families, the actin-dependent motor, myosin XI (a,b), and the putative chitin receptor Lyk5 (a,b,c). The APTi approach resulted in a homogenous population of transient P. patens mutants specific for our gene targets with zero surviving background plants within 8 d. The observed mutants directly corresponded to a maximal 93% reduction of myosin XI protein and complete loss of chitin-induced calcium spiking in the Lyk5-RNAi background. The positive selection nature of APTi represents a fundamental improvement in RNAi technology and will contribute to the growing demand for technologies amenable to high-throughput phenotyping.

Conditional genetic screen in Physcomitrella patens reveals a novel microtubule depolymerizing-end-tracking protein.

Our ability to identify genes that participate in cell growth and division is limited because their loss often leads to lethality. A solution to this is to isolate conditional mutants where the phenotype is visible under restrictive conditions. Here, we capitalize on the haploid growth-phase of the moss Physcomitrella patens to identify conditional loss-of-growth (CLoG) mutants with impaired growth at high temperature. We used whole-genome sequencing of pooled segregants to pinpoint the lesion of one of these mutants (clog1) and validated the identified mutation by rescuing the conditional phenotype by homologous recombination. We found that CLoG1 is a novel and ancient gene conserved in plants. At the restrictive temperature, clog1 plants have smaller cells but can complete cell division, indicating an important role of CLoG1 in cell growth, but not an essential role in cell division. Fluorescent protein fusions of CLoG1 indicate it is localized to microtubules with a bias towards depolymerizing microtubule ends. Silencing CLoG1 decreases microtubule dynamics, suggesting that CLoG1 plays a critical role in regulating microtubule dynamics. By discovering a novel gene critical for plant growth, our work demonstrates that P. patens is an excellent genetic system to study genes with a fundamental role in plant cell growth.

Chitin Triggers Calcium-Mediated Immune Response in the Plant Model Physcomitrella patens.

A characteristic feature of a plant immune response is the increase of the cytosolic calcium (Ca2+) concentration following infection, which results in the downstream activation of immune response regulators. The bryophyte Physcomitrella patens has been shown to mount an immune response when exposed to bacteria, fungi, or chitin elicitation, in a manner similar to the one observed in Arabidopsis thaliana. Nevertheless, whether the response of P. patens to microorganism exposure is Ca2+ mediated is currently unknown. Here, we show that P. patens plants treated with chitin oligosaccharides exhibit Ca2+ oscillations, and that a calcium ionophore can stimulate the expression of defense-related genes. Treatment with chitin oligosaccharides also results in an inhibition of growth, which can be explained by the depolymerization of the apical actin cytoskeleton of tip growing cells. These results suggest that chitin-triggered calcium oscillations are conserved and were likely present in the common ancestor of bryophytes and vascular plants.

F-Actin Mediated Focusing of Vesicles at the Cell Tip Is Essential for Polarized Growth.

F-actin has been shown to be essential for tip growth in an array of plant models, including Physcomitrella patens One hypothesis is that diffusion can transport secretory vesicles, while actin plays a regulatory role during secretion. Alternatively, it is possible that actin-based transport is necessary to overcome vesicle transport limitations to sustain secretion. Therefore, a quantitative analysis of diffusion, secretion kinetics, and cell geometry is necessary to clarify the role of actin in polarized growth. Using fluorescence recovery after photobleaching analysis, we first show that secretory vesicles move toward and accumulate at the tip in an actin-dependent manner. We then depolymerized F-actin to decouple vesicle diffusion from actin-mediated transport and measured the diffusion coefficient and concentration of vesicles. Using these values, we constructed a theoretical diffusion-based model for growth, demonstrating that with fast-enough vesicle fusion kinetics, diffusion could support normal cell growth rates. We further refined our model to explore how experimentally extrapolated vesicle fusion kinetics and the size of the secretion zone limit diffusion-based growth. This model predicts that diffusion-mediated growth is dependent on the size of the region of exocytosis at the tip and that diffusion-based growth would be significantly slower than normal cell growth. To further explore the size of the secretion zone, we used a cell wall degradation enzyme cocktail and determined that the secretion zone is smaller than 6 µm in diameter at the tip. Taken together, our results highlight the requirement for active transport in polarized growth and provide important insight into vesicle secretion during tip growth.

Characterization of Cell Boundary and Confocal Effects Improves Quantitative FRAP Analysis.

Fluorescence recovery after photobleaching (FRAP) is an important tool used by cell biologists to study the diffusion and binding kinetics of vesicles, proteins, and other molecules in the cytoplasm, nucleus, or cell membrane. Although many FRAP models have been developed over the past decades, the influence of the complex boundaries of 3D cellular geometries on the recovery curves, in conjunction with regions of interest and optical effects (imaging, photobleaching, photoswitching, and scanning), has not been well studied. Here, we developed a 3D computational model of the FRAP process that incorporates particle diffusion, cell boundary effects, and the optical properties of the scanning confocal microscope, and validated this model using the tip-growing cells of Physcomitrella patens. We then show how these cell boundary and optical effects confound the interpretation of FRAP recovery curves, including the number of dynamic states of a given fluorophore, in a wide range of cellular geometries-both in two and three dimensions-namely nuclei, filopodia, and lamellipodia of mammalian cells, and in cell types such as the budding yeast, Saccharomyces pombe, and tip-growing plant cells. We explored the performance of existing analytical and algorithmic FRAP models in these various cellular geometries, and determined that the VCell VirtualFRAP tool provides the best accuracy to measure diffusion coefficients. Our computational model is not limited only to these cells types, but can easily be extended to other cellular geometries via the graphical Java-based application we also provide. This particle-based simulation-called the Digital Confocal Microscopy Suite or DCMS-can also perform fluorescence dynamics assays, such as number and brightness, fluorescence correlation spectroscopy, and raster image correlation spectroscopy, and could help shape the way these techniques are interpreted.

Myosin XI localizes at the mitotic spindle and along the cell plate during plant cell division in Physcomitrella patens.

Cell division is a fundamental biological process that has been extensively investigated in different systems. Similar to most eukaryotic cells, plant cells assemble a mitotic spindle to separate replicated chromosomes. In contrast, to complete cell division, plant cells assemble a phragmoplast, which is composed of aligned microtubules and actin filaments. This structure helps transport vesicles containing new cell wall material, which then fuse to form the cell plate; the cell plate will expand to create the new dividing cell wall. Because vesicles are known to be transported by myosin motors during interphase, we hypothesized this could also be the case during cell division and we investigated the localization of the plant homologue of myosin V - myosin XI, in cell division. In this work, we used the protonemal cells of the moss Physcomitrella patens as a model, because of its simple cellular morphology and ease to generate transgenic cell lines expressing fluorescent tagged proteins. Using a fluorescent protein fusion of myosin XI, we found that, during mitosis, this molecule appears to associate with the kinetochores immediately after nuclear envelope breakdown. Following metaphase, myosin XI stays associated with the spindle's midzone during the rest of mitosis, and when the phragmoplast is formed, it concentrates at the cell plate. Using an actin polymerization inhibitor, latrunculin B, we found that the association of myosin XI with the mitotic spindle and the phragmoplast are only partially dependent on the presence of filamentous actin. We also showed that myosin XI on the spindle partially overlaps with a v-SNARE vesicle marker but is not co-localized with the endoplasmic reticulum and a RabA vesicle marker. These observations suggest an actin-dependent and an actin-independent behavior of myosin XI during cell division, and provide novel insights to our understanding of the function of myosin XI during plant cell division.

The kinesin-like proteins, KAC1/2, regulate actin dynamics underlying chloroplast light-avoidance in Physcomitrella patens.

In plants, light determines chloroplast position; these organelles show avoidance and accumulation responses in high and low fluence-rate light, respectively. Chloroplast motility in response to light is driven by cytoskeletal elements. The actin cytoskeleton mediates chloroplast photorelocation responses in Arabidopsis thaliana. In contrast, in the moss Physcomitrella patens, both, actin filaments and microtubules can transport chloroplasts. Because of the surprising evidence that two kinesin-like proteins (called KACs) are important for actin-dependent chloroplast photorelocation in vascular plants, we wanted to determine the cytoskeletal system responsible for the function of these proteins in moss. We performed gene-specific silencing using RNA interference in P. patens. We confirmed existing reports using gene knockouts, that PpKAC1 and PpKAC2 are required for chloroplast dispersion under uniform white light conditions, and that the two proteins are functionally equivalent. To address the specific cytoskeletal elements responsible for motility, this loss-of-function approach was combined with cytoskeleton-targeted drug studies. We found that, in P. patens, these KACs mediate the chloroplast light-avoidance response in an actin filament-dependent, rather than a microtubule-dependent manner. Using correlation-decay analysis of cytoskeletal dynamics, we found that PpKAC stabilizes cortical actin filaments, but has no effect on microtubule dynamics.

Apical myosin XI anticipates F-actin during polarized growth of Physcomitrella patens cells.

Tip growth is essential for land colonization by bryophytes, plant sexual reproduction and water and nutrient uptake. Because this specialized form of polarized cell growth requires both a dynamic actin cytoskeleton and active secretion, it has been proposed that the F-actin-associated motor myosin XI is essential for this process. Nevertheless, a spatial and temporal relationship between myosin XI and F-actin during tip growth is not known in any plant cell. Here, we use the highly polarized cells of the moss Physcomitrella patens to show that myosin XI and F-actin localize, in vivo, at the same apical domain and that both signals fluctuate. Surprisingly, phase analysis shows that increase in myosin XI anticipates that of F-actin; in contrast, myosin XI levels at the tip fluctuate in identical phase with a vesicle marker. Pharmacological analysis using a low concentration of the actin polymerization inhibitor latrunculin B showed that the F-actin at the tip can be significantly diminished while myosin XI remains elevated in this region, suggesting that a mechanism exists to cluster myosin XI-associated structures at the cell's apex. In addition, this approach uncovered a mechanism for actin polymerization-dependent motility in the moss cytoplasm, where myosin XI-associated structures seem to anticipate and organize the actin polymerization machinery. From our results, we inferred a model where the interaction between myosin XI-associated vesicular structures and F-actin polymerization-driven motility function at the cell's apex to maintain polarized cell growth. We hypothesize this is a general mechanism for the participation of myosin XI and F-actin in tip growing cells.

Actin interacting protein1 and actin depolymerizing factor drive rapid actin dynamics in Physcomitrella patens.

The remodeling of actin networks is required for a variety of cellular processes in eukaryotes. In plants, several actin binding proteins have been implicated in remodeling cortical actin filaments (F-actin). However, the extent to which these proteins support F-actin dynamics in planta has not been tested. Using reverse genetics, complementation analyses, and cell biological approaches, we assessed the in vivo function of two actin turnover proteins: actin interacting protein1 (AIP1) and actin depolymerizing factor (ADF). We report that AIP1 is a single-copy gene in the moss Physcomitrella patens. AIP1 knockout plants are viable but have reduced expansion of tip-growing cells. AIP1 is diffusely cytosolic and functions in a common genetic pathway with ADF to promote tip growth. Specifically, ADF can partially compensate for loss of AIP1, and AIP1 requires ADF for function. Consistent with a role in actin remodeling, AIP1 knockout lines accumulate F-actin bundles, have fewer dynamic ends, and have reduced severing frequency. Importantly, we demonstrate that AIP1 promotes and ADF is essential for cortical F-actin dynamics.

Myosin XI is essential for tip growth in Physcomitrella patens.

Class XI myosins are plant specific and responsible for cytoplasmic streaming. Because of the large number of myosin XI genes in angiosperms, it has been difficult to determine their precise role, particularly with respect to tip growth. The moss Physcomitrella patens provides an ideal system to study myosin XI function. P. patens has only two myosin XI genes, and these genes encode proteins that are 94% identical to each other. To determine their role in tip growth, we used RNA interference to specifically silence each myosin XI gene using 5' untranslated region sequences. We discovered that the two myosin XI genes are functionally redundant, since silencing of either gene does not affect growth or polarity. However, simultaneous silencing of both myosin XIs results in severely stunted plants composed of small rounded cells. Although similar to the phenotype resulting from silencing of other actin-associated proteins, we show that this phenotype is not due to altered actin dynamics. Consistent with a role in tip growth, we show that a functional, full-length fusion of monomeric enhanced green fluorescent protein (mEGFP) to myosin XI accumulates at a subcortical, apical region of actively growing protonemal cells.

Quantitative analysis of organelle distribution and dynamics in Physcomitrella patens protonemal cells.

BACKGROUND: In the last decade, the moss Physcomitrella patens has emerged as a powerful plant model system, amenable for genetic manipulations not possible in any other plant. This moss is particularly well suited for plant polarized cell growth studies, as in its protonemal phase, expansion is restricted to the tip of its cells. Based on pollen tube and root hair studies, it is well known that tip growth requires active secretion and high polarization of the cellular components. However, such information is still missing in Physcomitrella patens. To gain insight into the mechanisms underlying the participation of organelle organization in tip growth, it is essential to determine the distribution and the dynamics of the organelles in moss cells. RESULTS: We used fluorescent protein fusions to visualize and track Golgi dictyosomes, mitochondria, and peroxisomes in live protonemal cells. We also visualized and tracked chloroplasts based on chlorophyll auto-fluorescence. We showed that in protonemata all four organelles are distributed in a gradient from the tip of the apical cell to the base of the sub-apical cell. For example, the density of Golgi dictyosomes is 4.7 and 3.4 times higher at the tip than at the base in caulonemata and chloronemata respectively. While Golgi stacks are concentrated at the extreme tip of the caulonemata, chloroplasts and peroxisomes are totally excluded. Interestingly, caulonemata, which grow faster than chloronemata, also contain significantly more Golgi dictyosomes and fewer chloroplasts than chloronemata. Moreover, the motility analysis revealed that organelles in protonemata move with low persistency and average instantaneous speeds ranging from 29 to 75 nm/s, which are at least three orders of magnitude slower than those of pollen tube or root hair organelles. CONCLUSIONS: To our knowledge, this study reports the first quantitative analysis of organelles in Physcomitrella patens and will make possible comparisons of the distribution and dynamics of organelles from different tip growing plant cells, thus enhancing our understanding of the mechanisms of plant polarized cell growth.

Exocytosis precedes and predicts the increase in growth in oscillating pollen tubes.

We examined exocytosis during oscillatory growth in lily (Lilium formosanum and Lilium longiflorum) and tobacco (Nicotiana tabacum) pollen tubes using three markers: (1) changes in cell wall thickness by Nomarski differential interference contrast (DIC), (2) changes in apical cell wall fluorescence in cells stained with propidium iodide (PI), and (3) changes in apical wall fluorescence in cells expressing tobacco pectin methyl esterase fused to green fluorescent protein (PME-GFP). Using PI fluorescence, we quantified oscillatory changes in the amount of wall material from both lily and tobacco pollen tubes. Measurement of wall thickness by DIC was only possible with lily due to limitations of microscope resolution. PME-GFP, a direct marker for exocytosis, only provides information in tobacco because its expression in lily causes growth inhibition and cell death. We show that exocytosis in pollen tubes oscillates and leads the increase in growth rate; the mean phase difference between exocytosis and growth is -98 degrees +/- 3 degrees in lily and -124 degrees +/- 4 degrees in tobacco. Statistical analyses reveal that the anticipatory increase in wall material predicts, to a high degree, the rate and extent of the subsequent growth surge. Exocytosis emerges as a prime candidate for the initiation and regulation of oscillatory pollen tube growth.

Rapid formin-mediated actin-filament elongation is essential for polarized plant cell growth.

Formins are present in all eukaryotes and are essential for the creation of actin-based structures responsible for diverse cellular processes. Because multicellular organisms contain large formin gene families, establishing the physiological functions of formin isoforms has been difficult. Using RNAi, we analyzed the function of all 9 formin genes within the moss Physcomitrella patens. We show that plants lacking class II formins (For2) are severely stunted and composed of spherical cells with disrupted actin organization. In contrast, silencing of all other formins results in normal elongated cell morphology and actin organization. Consistent with a role in polarized growth, For2 are apically localized in growing cells. We show that an N-terminal phosphatase tensin (PTEN)-like domain mediates apical localization. The PTEN-like domain is followed by a conserved formin homology (FH)1-FH2 domain, known to promote actin polymerization. To determine whether apical localization of any FH1-FH2 domain mediates polarized growth, we performed domain swapping. We found that only the class II FH1-FH2, in combination with the PTEN-like domain, rescues polarized growth, because it cannot be replaced with a similar domain from a For1. We used in vitro polymerization assays to dissect the functional differences between these FH1-FH2 domains. We found that both the FH1 and the FH2 domains from For2 are required to mediate exceptionally rapid rates of actin filament elongation, much faster than any other known formin. Thus, our data demonstrate that rapid rates of actin elongation are critical for driving the formation of apical filamentous actin necessary for polarized growth.