Synthetic biology as a tool to online teaching undergraduate level molecular biology.

During the Covid-19 pandemic, all on-campus activities were required to be adapted to a distance-learning format, which was especially challenging for practical courses. Molecular biology is a mandatory course for biology undergraduates at Universidade Federal Fluminense. To overcome the impossibility of practical activities, we proposed problem-solving synthetic biology assignments to review and consolidate the knowledge acquired in the molecular biology course. A survey indicated that the synthetic biology challenges were effective to promote students' active learning and worked as a proper substitute to the practical classes.

The plant cell cycle: Pre-Replication complex formation and controls.

The multiplication of cells in all living organisms requires a tight regulation of DNA replication. Several mechanisms take place to ensure that the DNA is replicated faithfully and just once per cell cycle in order to originate through mitoses two new daughter cells that contain exactly the same information from the previous one. A key control mechanism that occurs before cells enter S phase is the formation of a pre-replication complex (pre-RC) that is assembled at replication origins by the sequential association of the origin recognition complex, followed by Cdt1, Cdc6 and finally MCMs, licensing DNA to start replication. The identification of pre-RC members in all animal and plant species shows that this complex is conserved in eukaryotes and, more importantly, the differences between kingdoms might reflect their divergence in strategies on cell cycle regulation, as it must be integrated and adapted to the niche, ecosystem, and the organism peculiarities. Here, we provide an overview of the knowledge generated so far on the formation and the developmental controls of the pre-RC mechanism in plants, analyzing some particular aspects in comparison to other eukaryotes.

The plant cell cycle: Pre-Replication complex formation and controls

Abstract The multiplication of cells in all living organisms requires a tight regulation of DNA replication. Several mechanisms take place to ensure that the DNA is replicated faithfully and just once per cell cycle in order to originate through mitoses two new daughter cells that contain exactly the same information from the previous one. A key control mechanism that occurs before cells enter S phase is the formation of a pre-replication complex (pre-RC) that is assembled at replication origins by the sequential association of the origin recognition complex, followed by Cdt1, Cdc6 and finally MCMs, licensing DNA to start replication. The identification of pre-RC members in all animal and plant species shows that this complex is conserved in eukaryotes and, more importantly, the differences between kingdoms might reflect their divergence in strategies on cell cycle regulation, as it must be integrated and adapted to the niche, ecosystem, and the organism peculiarities. Here, we provide an overview of the knowledge generated so far on the formation and the developmental controls of the pre-RC mechanism in plants, analyzing some particular aspects in comparison to other eukaryotes.

AIP1 is a novel Agenet/Tudor domain protein from Arabidopsis that interacts with regulators of DNA replication, transcription and chromatin remodeling.

BACKGROUND: DNA replication and transcription are dynamic processes regulating plant development that are dependent on the chromatin accessibility. Proteins belonging to the Agenet/Tudor domain family are known as histone modification "readers" and classified as chromatin remodeling proteins. Histone modifications and chromatin remodeling have profound effects on gene expression as well as on DNA replication, but how these processes are integrated has not been completely elucidated. It is clear that members of the Agenet/Tudor family are important regulators of development playing roles not well known in plants. METHODS: Bioinformatics and phylogenetic analyses of the Agenet/Tudor Family domain in the plant kingdom were carried out with sequences from available complete genomes databases. 3D structure predictions of Agenet/Tudor domains were calculated by I-TASSER server. Protein interactions were tested in two-hybrid, GST pulldown, semi-in vivo pulldown and Tandem Affinity Purification assays. Gene function was studied in a T-DNA insertion GABI-line. RESULTS: In the present work we analyzed the family of Agenet/Tudor domain proteins in the plant kingdom and we mapped the organization of this family throughout plant evolution. Furthermore, we characterized a member from Arabidopsis thaliana named AIP1 that harbors Agenet/Tudor and DUF724 domains. AIP1 interacts with ABAP1, a plant regulator of DNA replication licensing and gene transcription, with a plant histone modification "reader" (LHP1) and with non modified histones. AIP1 is expressed in reproductive tissues and its down-regulation delays flower development timing. Also, expression of ABAP1 and LHP1 target genes were repressed in flower buds of plants with reduced levels of AIP1. CONCLUSIONS: AIP1 is a novel Agenet/Tudor domain protein in plants that could act as a link between DNA replication, transcription and chromatin remodeling during flower development.

ABAP1 is a novel plant Armadillo BTB protein involved in DNA replication and transcription.

In multicellular organisms, organogenesis requires a tight control of the balance between cell division and cell differentiation. Distinct signalling pathways that connect both cellular processes with developmental cues might have evolved to suit different developmental plans. Here, we identified and characterized a novel protein that interacts with pre-replication complex (pre-RC) subunits, designated Armadillo BTB Arabidopsis protein 1 (ABAP1). Overexpression of ABAP1 in plants limited mitotic DNA replication and decreased cell proliferation in leaves, whereas ABAP1 downregulation increased cell division rates. Activity of ABAP1 in transcription was supported by its association with the transcription factor AtTCP24. The ABAP1-AtTCP24 complex bound specifically to the promoters of AtCDT1a and AtCDT1b in vitro and in vivo. Moreover, expression levels of AtCDT1a and AtCDT1b were reduced in ABAP1-overexpressing plants and they were increased in plants with reduced levels of ABAP1. We propose that ABAP1 participates in a negative feedback loop regulating mitotic DNA replication during leaf development, either by repressing transcription of pre-RC genes and possibly by regulating pre-RC utilization through direct association with pre-RC components.