Retinoblastoma protein regulates cell proliferation, differentiation, and endoreduplication in plants

Abstract

SummaryRetinoblastoma protein (Rb) plays a key role in cell cycle control, cell differentiation, and apoptosis in animals. In this study, we used virus-induced gene silencing (VIGS) to investigate the cellular functions of Rb in higher plants. VIGS of NbRBR1, which encodes the Nicotiana benthamiana Rb homolog, resulted in growth retardation and abnormal organ development. At the cellular level, Rb suppression caused prolonged cell proliferation in tissues that are normally differentiated, which indicates that Rb is a negative regulator of plant cell division. Furthermore, differentiation of the epidermal pavement cells and trichomes was partially retarded, and stomatal clusters formed in the epidermis, likely due to uncontrolled cell division of stomata precursor cells. Rb suppression also caused extra DNA replication in endoreduplicating leaf cells, suggesting a role of Rb in the endocycle. These Rb phenotypes were accompanied by stimulated transcription of E2F and E2F-regulated S-phase genes. Thus, disruption of Rb function in plants leads to ectopic cell division in major organs that correlates with a delay in cell differentiation as well as increased endoreduplication, which indicates that Rb coordinates these processes in plant organ development.

Divide and conquer: development and cell cycle genes in plant transformation

“…To aid recognition of meristemoids at early stages in development, the use of transgenic marker genotypes, such as those containing a WUSCHEL-reporter or STM-reporter construct [58,59], would be useful. In addition, monitoring of S-phase-specific genes, such as RNR, MCM, CDC6, PCNA, H2A, H4 and E2Fa [23][24][25]38], might give detailed insights into the mechanisms of cell cycle modulation: using microarray analysis, O100 genes related to S phase have been identified in Arabidopsis after overexpression of E2Fa-DPa [24]. In addition, changes in heterochromatin remodeling, DNA or histone methylation and histone acetylation and deacetylation can be directly monitored by a variety of methods [17][18][19]60].…”

“…Experimentally, virus-induced gene silencing (VIGS) of RBR in tobacco resulted in the up-regulated transcription of E2F and S phase active genes, such as ribonucleotide reductase (RNR), proliferating cell nuclear antigen (PCNA), minichromosome maintenance (MCM), histone H1 and replication origin protein (CDC6), in addition to an increased proportion of 4C nuclei [38]. Calreticulins and calpains can regulate regeneration Fluctuations of Ca 2C concentration in subcellular compartments can act as a signal of cell stress, in communication between the plasma membrane and organelles, and in the control of the cell cycle [39].…”

“…However, it is also abundantly clear that their constitutive activation or silencing would prevent normal development. Published examples include suppression of the RBR protein by VIGS (leading to ectopic cell division and a lack of differentiation) [38], suppression of phytocalpain NbDEK1 (inhibiting cell differentiation) [43] and overexpression of the calreticulin BrCRT1 (retarding plant growth) [42]. There are several inducible systems available for the spatial and temporal control of gene expression.…”

Grasses Like Mammals? Redundancy and Compensatory Regulation within the Retinoblastoma Protein Family

“…This context-dependent functional complementation between pocket proteins provides a basis to explain the emergence of RB-dependent sporadic tumors in senescent tissues. Thus far, RBR inactivation has been reported in plant species with apparently a single RBR gene, such as Arabidopsis 14 and Nicotiana, 15 or in maize undifferentiated cells. 11 It would be interesting to know whether a differential response of naive and differentiated/senescent cells to the loss of function of one pocket protein, similar to that identified in mammals, is also true for plants, such as maize, with a complex and interdependent RBR family of proteins.…”

Regulating DNA Replication in Plants

“…CDKA/CYCD3 are responsible for the phosphorylation of RBR (RETINOBLASTOMA-RELATED), the homolog of the mammalian RB protein. In fact, the RBR pathway regulates the number of endocycles once cells have exited the cell cycle (Park et al 2005;Desvoyes et al 2006;Borghi et al 2010). Whether it is a direct effect or mediated by downstream RBR/E2F targets is not clear.…”