Biology subjects

Tolmie, A. F.

Publications and source records attributed to Tolmie, A. F..

3 recordsLinked to original sources

bioRxiv2019

The Maize LINC KASH AtSINE-like2 (MLKS2) gene encodes an ARM domain protein that tethers the nucleus to F-actin and is required for normal development and meiotic chromosome segregation

The linker of nucleoskeleton and cytoskeleton (LINC) complex is an essential multi-protein structure spanning the eukaryotic nuclear envelope. The LINC complex functions to maintain nuclear architecture, positioning, and mobility, along with specialized functions in meiotic prophase and chromosome segregation. Members of the LINC complex were recently identified in maize, an important scientific and agricultural grass species. Here we characterized Maize LINC KASH AtSINE-like2, MLKS2, which encodes a highly conserved SINE-group plant KASH protein with characteristic N-terminal armadillo repeats (ARM). Using a heterologous expression system, we showed that actively expressed GFP-MLKS2 is targeted to the nuclear periphery and colocalizes with F-actin and the endoplasmic reticulum, but not microtubules in the cell cortex. Expression of GFP-MLKS2, but not GFP-MLKS2{Delta}ARM, resulted in nuclear anchoring. Genetic analysis of transposon-insertion mutations, mlks2-1 and mlks2-2, showed that the mutant phenotypes were pleiotropic, affecting root hair nuclear morphology, stomatal complex development, multiple aspects of meiosis, and pollen viability. In male meiosis, the mutants showed defects for bouquet-stage telomere clustering, nuclear repositioning, perinuclear actin accumulation, dispersal of late prophase bivalents, and meiotic chromosome segregation. These findings support a model in which the nucleus is connected to cytoskeletal F-actin through the ARM-domain, predicted alpha solenoid structure of MLKS2. Functional conservation of MLKS2 was demonstrated through genetic rescue of the misshapen nuclear phenotype of an Arabidopsis (triple-WIP) KASH mutant. This study establishes a role for the SINE-type KASH proteins in affecting the dynamic nuclear phenomena required for normal plant growth and fertility.

cell biology↗

bioRxiv2018

The cell wall regulates dynamics and size of plasma-membrane nanodomains in Arabidopsis.

Plant plasma-membrane (PM) proteins are involved in several vital processes, such as detection of pathogens, solute transport and cellular signalling. Recent models suggest that for these proteins to function effectively there needs to be structure within the PM allowing, for example, proteins in the same signalling cascade to be spatially organized. Here we demonstrate that several proteins with divergent functions are located in clusters of differing size in the membrane using sub-diffraction-limited Airyscan confocal microscopy. In addition, single particle tracking reveals that these proteins move at different rates within the membrane. We show that the actin and microtubule cytoskeletons appear to significantly regulate the mobility of one of these proteins (the pathogen receptor FLS2) and we further demonstrate that the cell wall is critical for the regulation of cluster size by affecting single particle dynamics of two proteins with key roles in morphogenesis (PIN3) and pathogen perception (FLS2). We propose a model in which the cell wall and cytoskeleton are pivotal for differentially regulating protein cluster size and dynamics thereby contributing to the formation and functionality of membrane nanodomains.\n\nOne sentence summarySize and mobility of protein nanodomains in the plant plasma-membrane are regulated by interaction with the cell wall extracellular matrix.\n\nSignificance statementThe plant plasma membrane acts as the front line for cellular perception of the environment. As such, a large number of signalling and transport proteins which perceive or transport environmental signals, developmental cues and nutrients are located within it. Recently, a number of studies have revealed that proteins located within the plasma membrane do not simply freely diffuse within its plane. Rather, proteins are localized in nanometer sized structures called nanodomains. In addition to the plasma-membrane, plant cells also have an extracellular matrix, the cell wall. Here we have shown that the cell wall has a role in regulating the dynamics and size of plasma membrane nanodomains for proteins involved in morphogenesis (PIN3) and pathogen perception (FLS2).

cell biology↗

bioRxiv2018

Identification and characterization of conserved and divergent genes encoding the nuclear envelope LINC complex in maize (Zea mays L.).

The LINC (Linker of Nucleoskeleton to Cytoskeleton) complex is an essential multi-protein structure spanning the nuclear envelope. It connects the cytoplasm to the nucleoplasm, functions to maintain nuclear shape and architecture, and regulates chromosome dynamics during cell division. Knowledge of LINC complex composition and function in the plant kingdom is primarily limited to Arabidopsis, but critically missing from the evolutionarily distant monocots which include grasses, the most important agronomic crops worldwide. To fill this knowledge gap, we identified and characterized 22 maize genes, including a new grass-specific KASH gene family. Using bioinformatic, biochemical, and cell biological approaches, we provide evidence that representative KASH candidates localize to the nuclear periphery and interact with ZmSUN2 in vivo. FRAP experiments using domain-deletion constructs verified that this SUN-KASH interaction was dependent on the SUN but not the coiled-coil domain of ZmSUN2. A summary working model is proposed for the entire maize LINC complex encoded by conserved and divergent gene families. These findings expand our knowledge of the plant nuclear envelope in a model grass species, with implications for both basic and applied cellular research.\n\nSUMMARY STATEMENTGenes encoding maize candidates for the core LINC and associated complex proteins have been comprehensively identified with functional validation by one or more assays for several of the KASH genes.

cell biology↗