The cell-cycle promoter cdc2aAt from Arabidopsis thaliana is induced in the lateral roots of the actinorhizal tree Allocasuarina verticillata during the early stages of the symbiotic interaction with Frankia

The symbiosis between the actinorhizal tree Allocasuarina verticillata and the actinomycete Frankia leads to the formation of root nodules inside which bacteria fix atmospheric nitrogen. Actinorhizal nodule organogenesis starts with the induction of cell divisions in the root cortex and in the pericycle cells opposite protoxylem poles near Frankia -infected root hairs. To study the ability of Frankia to induce progression through the cell cycle, we monitored the expression of the β-glucuronidase ( gus ) gene driven by the promoter from cdc2aAt , an Arabidopsis cyclin-dependent kinase gene that displays competence for cell division, during plant growth and nodule ontogenesis. In non-symbiotic tissues, the gus gene was mainly expressed in primary and secondary meristems of roots and shoots. Auxins and cytokinins were found to induce reporter gene activity in the root system of whole plants, showing that the promoter cdc2aAt displayed the same regulation by hormones in Allocasuarina as that reported in Arabidopsis . In transgenic nodules, gus expression was found to be restricted to the phellogen. During the early stages of the interaction between Frankia and the plant root system, cdc2aAt was strongly induced in the lateral roots surrounded by hyphae of the actinomycete. Histochemical analysis of β-glucuronidase activity revealed that cells from the pericycle opposite protoxylem poles were very deeply stained. These data indicate that upon Frankia infection, cells from the lateral roots, and notably pericycle cells that can give rise to a nodule or a root primordium, prepare to re-enter the cell cycle.     

Increased leakiness of the tetracycline-inducible Triple-Op promoter in dividing cells renders it unsuitable for high inducible levels of a dominant negative CDC2aAt gene

A tetracycline-inducible promoter system was used to generate transgenic tobacco plants that confer inducible expression of the wild type or a dominant negative allele of the gene coding for the cyclin-dependent kinase (CDK) of Arabidopsis thaliana CDC2aAt. Although the total extractable CDK activity was doubled, the induced expression of the wild-type CDC2aAt did not correlate with any change of the cell cycle kinetics. An increase of CDK activity upon CDC2aAt expression was only seen in dividing cell populations, demonstrating that CDC2aAt expression itself is not sufficient to induce CDK activation. Induced expression of the dominant negative CDC2aAt.N146 correlated with a reduction of CDK activity to 66% of the level found in non-induced cells. This decrease was not sufficient to block cell division. The isolation of plants showing only low inducible levels of CDC2aAt.N146 suggests that a counterselection against strong inducible lines had occurred. Accordingly, Triple-Op promoter activity was found in dividing cells in the absence of tetracycline.     

Alfin1 transcription factor overexpression enhances plant root growth under normal and saline conditions and improves salt tolerance in alfalfa

Plant root development is an essential determinant of plant growth and crop yield that could be enhanced by induced changes in the expression of root-specific regulatory factors. We reported previously that Alfin1 binds DNA in a sequence-specific manner and that Alfin1 overexpression in transgenic alfalfa (Medicago sativa L.) enhances expression of the salt-inducible MsPRP2 gene in roots, suggesting that Alfin1 functions to regulate gene expression in roots. Here we show that Alfin1 is an essential gene for root growth and that its overexpression in transgenic plants confers a many-fold increase in root growth under normal and saline conditions. Alfin1-binding sites occur in promoters of genes expressed in roots of a wide variety of plant species and we propose that it is a general root growth regulator. Even though Alfin1 overexpression was under the control of the CaMV 35S promoter, plant shoot growth was not adversely affected. We show further that introduction of the Alfin1 transgene in plants confers a dominant characteristic that significantly increases plant growth and salt tolerance.     

The STT3a subunit isoform of the Arabidopsis oligosaccharyltransferase controls adaptive responses to salt/osmotic stress

Arabidopsis stt3a-1 and stt3a-2 mutations cause NaCl/osmotic sensitivity that is characterized by reduced cell division in the root meristem. Sequence comparison of the STT3a gene identified a yeast ortholog, STT3, which encodes an essential subunit of the oligosaccharyltransferase complex that is involved in protein N-glycosylation. NaCl induces the unfolded protein response in the endoplasmic reticulum (ER) and cell cycle arrest in root tip cells of stt3a seedlings, as determined by expression profiling of ER stress-responsive chaperone (BiP-GUS) and cell division (CycB1;1-GUS) genes, respectively. Together, these results indicate that plant salt stress adaptation involves ER stress signal regulation of cell cycle progression. Interestingly, a mutation (stt3b-1) in another Arabidopsis STT3 isogene (STT3b) does not cause NaCl sensitivity. However, the stt3a-1 stt3b-1 double mutation is gametophytic lethal. Apparently, STT3a and STT3b have overlapping and essential functions in plant growth and developmental processes, but the pivotal and specific protein glycosylation that is a necessary for recovery from the unfolded protein response and for cell cycle progression during salt/osmotic stress recovery is associated uniquely with the function of the STT3a isoform.     

Tuberin negatively affects BCL-2’s cell survival function

Summary. Uncontrolled cell cycle progression and cell growth are key properties of tumor cells. The tumor suppressor genes responsible for the autosomal dominantly inherited disease tuberous sclerosis (TSC) have been demonstrated to control both, cell cycle and cell size regulation. Hamartin, encoded by TSC1, and tuberin, encoded by TSC2, form a complex, of which tuberin is assumed to be the functional component. Loss of TSC genes function triggers hamartoma development in TSC patients. However, in vivo mostly tumor cell development is rapidly terminated via apoptosis. BCL-2, the founding member of the BCL-2 family of proteins, is well known for its anti-apoptotic properties. Here we show that pro-apoptotic actinomycin D cannot interfere with BCL-2’s cell survival functions. However, we found tuberin to negatively regulate BCL-2’s anti-apoptotic effects on low serum-induced apoptosis. These findings warrant further investigations to elucidate the molecular mechanism underlying tuberin’s negative effects on cell survival.     

The division of Drosophila germline stem cells and their precursors requires a specific cyclin

A fundamental yet essentially unexplored question in stem cell biology is whether the stem cell cycle has specific features. Three B-cyclins in Drosophila, Cyclins (Cyc) A, B, and B3, associate with CDK1 and play partially redundant roles in embryogenic mitosis . Here, we show that the division of Drosophila GSCs and their precursors, the primordial germ cells (PGCs), specifically requires CycB. CycB is ubiquitously expressed in both germline and somatic lineages. However, CycB mutation does not have obvious effect on somatic development but causes PGCs to severely under proliferate. Moreover, both female and male CycB mutant GSCs fail to be maintained properly. Removing Cyclin B specifically from female GSCs causes the same defect, confirming the direct and cell-autonomous function of Cyclin B for GSC division. In contrast, two other G2 cyclins, CycA and CycB3, are also expressed in PGCs and GSCs, but overexpressing CycA cannot rescue the CycB mutant defects. These results indicate that the requirement of CycB for PGC and GSC divisions unlikely reflects the insufficient level of G2 cyclins in the CycB mutant but is in favor of a distinct function of CycB in these cells. Our results indicate that stem cells may use specific cell cycle regulators for their division.     

Competency of Nicotiana tabacum L. stem tissues to dedifferentiate is associated with differential levels of cell cycle gene expression and endogenous cytokinins

The expression of the mitotic cyclin Arath; CYCB1;1 and of the cyclin-dependent kinase Arath; CDC2a was located by beta-glucuronidase histochemical detection and in situ hybridization, and was quantified by 4-methylumbelliferyl beta- D-glucuronide assay in tobacco stem tissues during both in vivo differentiation and in vitro dedifferentiation. The changes in localization of endogenous cytokinins were also determined during both processes using quantitative analysis and in situ immunocytochemistry. The CDC2a promoter was expressed continuously during stem development, with particularly high expression in the shoot apical meristem and in the internal and external primary phloem. CYCB1 expression was not restricted to the dividing cells; its expression in the shoot apical meristem was particularly high in the leaf-forming peripheral cells but the gene was also expressed throughout development in the internal and external phloem in which the rate of cell division was reduced or zero. Following in vitro culture, the internal phloem cells appeared to be particularly competent to re-enter the cell cycle within a short lag phase while the pith tissue reactivated later. In culture, cells that resumed division were found to accumulate cytokinins. The high competency of primary phloem to dedifferentiate was associated with its capacity to express CDC2a and CYCB genes and the presence of high cytokinin levels, providing some insights into the determinants of competency for resuming cell division.