SCL28 promotes cell expansion and endoreplication in Arabidopsis by activating SIAMESE-RELATED cyclin-dependent kinase inhibitors.

The processes that contribute to plant organ morphogenesis are spatial-temporally organized. Within the meristem the mitotic cell cycle produces new cells that subsequently engage in specific cell expansion and differentiation programs once they exit the division competent zone. The latter is frequently accompanied by endoreplication, being an alternative cell cycle that replicates the DNA without nuclear division, causing a stepwise increase in somatic ploidy. We have previously shown that the Arabidopsis SCL28 transcription factor promotes progression through G2/M and modulates division plane orientation. Here, we demonstrate that SCL28 co-express and regulates genes specific to cell elongation and differentiation, including genes related to cell wall and cytoskeleton assembly. Consistently, this correlates with defects in post-mitotic cell expansion in a scl28 mutant. Strikingly, SCL28 controls expression of 6 members of the SIAMESE/SIAMESE-RELATED (SIM/SMR) family, encoding cyclin-dependent kinase inhibitors with a role in promoting mitotic cell cycle exit and endoreplication onset, both in response to developmental and environmental cues. Consistent with this role, scl28 mutants displayed reduced endoreplication, both in roots and leaves. Altogether, these results suggest that SCL28 controls cell expansion and differentiation by promoting endoreplication onset and by modulating aspects of the biogenesis, assembly and remodeling of the cytoskeleton and cell wall.

A quantitative gibberellin signalling biosensor reveals a role for gibberellins in internode specification at the shoot apical meristem

Growth at the shoot apical meristem (SAM) is essential for shoot architecture construction. The phytohormones gibberellins (GA) play a pivotal role in coordinating plant growth, but their role in the SAM remains mostly unknown. Here, we developed a ratiometric GA signalling biosensor by engineering one of the DELLA repressors, to suppress its master regulatory function in GA transcriptional responses while preserving its degradation upon GA sensing. We demonstrate that this novel degradation-based biosensor accurately reports on cellular changes in GA levels and perception during development. We used this biosensor to map GA signalling activity in the SAM. We show that high GA signalling is found primarily in cells located between organ primordia that are the precursors of internodes. By gain- and loss-of-function approaches, we further demonstrate that GAs regulate cell division plane orientation to establish the typical cellular organisation of internodes, thus contributing to internode specification in the SAM.

Functional characterization of TANGLED1 interaction with PHRAGMOPLAST ORIENTING KINESIN1 during mitosis in Arabidopsis

Cell division requires spatial coordination to properly position the division plane. How division plane positioning contributes to plant growth remains unknown. Two unrelated microtubule binding proteins, TANGLED1 (TAN1) and AUXIN-INDUCED-IN-ROOT-CULTURES9 (AIR9), are together required for normal Arabidopsis growth and division. tan1 air9 double mutants have synthetic growth and division plane orientation defects while single mutants lack obvious defects. We show that the first 132 amino acids of TAN1 (TAN1(1-132)) rescue the tan1 air9 double mutant and localize to the division site during telophase. Loss of both rescue and division-site localization occurred when interaction between TAN1 and PHRAGMOPLAST ORIENTING KINESIN1 (POK1) was disrupted by replacing six amino acid residues with alanines in TAN1(1-132). However, full-length TAN1 with the same alanine substitutions significantly rescued the tan1 air9 double mutant and remained at the division site throughout mitosis, although its accumulation was reduced and phragmoplast positioning defects occurred. POK1 often fails to accumulate at the division site in tan1 air9 mutants, suggesting that both TAN1 and AIR9 stabilize POK1 there. Finally, a mitosis specific promoter driving TAN1 rescued the tan1 air9 double mutant phenotypes indicating that defects seen in the root differentiation zone reflect the loss of mitotic-specific TAN1 activity.

SABRE populates ER domains essential for cell plate maturation and cell expansion influencing cell and tissue patterning

SABRE, which is found throughout eukaryotes and was originally identified in plants, mediates cell expansion, division plane orientation, and planar polarity in plants. How and where SABRE mediates these processes remain open questions. We deletedSABREinPhyscomitrium patens, an excellent model for cell biology.SABREnull mutants were stunted, similar to phenotypes in seed plants. Additionally, polarized growing cells were delayed in cytokinesis, sometimes resulting in catastrophic failures. A functional SABRE fluorescent fusion protein localized to dynamic puncta on regions of the endoplasmic reticulum (ER) during interphase and at the cell plate during cell division. WithoutSABRE, cells accumulated ER aggregates and the ER abnormally buckled along the developing cell plate. Notably, callose deposition was delayed in∆sabre, and in cells that failed to divide, abnormal callose accumulations formed at the cell plate. Our findings revealed a surprising and fundamental role for the ER in cell plate maturation.

TANGLED1 mediates microtubule interactions that may promote division plane positioning in maize

The microtubule cytoskeleton serves as a dynamic structural framework for mitosis in eukaryotic cells. TANGLED1 (TAN1) is a microtubule-binding protein that localizes to the division site and mitotic microtubules and plays a critical role in division plane orientation in plants. Here, in vitro experiments demonstrate that TAN1 directly binds microtubules, mediating microtubule zippering or end-on microtubule interactions, depending on their contact angle. Maize tan1 mutant cells improperly position the preprophase band (PPB), which predicts the future division site. However, cell shape–based modeling indicates that PPB positioning defects are likely a consequence of abnormal cell shapes and not due to TAN1 absence. In telophase, colocalization of growing microtubules ends from the phragmoplast with TAN1 at the division site suggests that TAN1 interacts with microtubule tips end-on. Together, our results suggest that TAN1 contributes to microtubule organization to ensure proper division plane orientation.

TANGLED1 mediates interactions between microtubules that may promote spindle organization and phragmoplast guidance to the division site in maize

The microtubule cytoskeleton serves as a dynamic structural framework for mitosis in eukaryotic cells. TANGLED1 (TAN1) is a microtubule-binding protein that localizes to the division site and mitotic microtubules and plays a critical role in division plane orientation in plants. Here, in vitro experiments demonstrate that TAN1 directly binds microtubules, mediating microtubule zippering or end-on microtubule interactions, depending on their contact angle. Maize tan1 mutant cells improperly position the preprophase band (PPB), which predicts the future division site. However, cell-shape-based modeling indicates that PPB positioning defects are likely a consequence of abnormal cell shapes and not due to TAN1 absence. Spindle defects in the tan1 mutant suggest that TAN1-mediated microtubule zippering may contribute to metaphase spindle organization. In telophase, co-localization of growing microtubules ends from the phragmoplast with TAN1 at the division site suggests that TAN1 interacts with microtubule tips end-on. Together, our results suggest that TAN1 contributes to spindle and phragmoplast microtubule organization to ensure proper division plane orientation.

Mechanosensitive nuclear asymmetries define a bipolar spindle scaffold to ensure mitotic fidelity

During prophase, centrosomes need to separate and position to correctly assemble the mitotic spindle. This process occurs through the action of molecular motors, cytoskeletal networks and the nucleus. How the combined activity of these different components is spatiotemporally regulated to ensure efficient spindle assembly remains unclear. Here we show that during prophase the centrosomes-nucleus axis reorients, so that centrosomes are positioned on the shortest nuclear axis at nuclear envelope (NE) breakdown. This centrosomes-nucleus configuration depends on mechanical cues generated by mitotic chromosome condensation on the prophase nucleus. We further show these mechanosensitive cues act through SUN1/2 and NudE+NudEL to enable the polarized loading of Dynein on the NE. Finally, we observe this centrosome configuration favors the establishment of an initial bipolar spindle scaffold, facilitating chromosome capture and accurate segregation, without compromising division plane orientation. We propose that chromosome segregation fidelity depends on the mechanical properties of the prophase nucleus that facilitate spindle assembly by regulating NE-Dynein localization.