A Cassava CPRF-2-like bZIP Transcription Factor Showed Increased Transcript Levels during Light Treatment
Authors: Pontes Lígia Cristine Gonçalves , Cardoso Cristina Michiko Yokoyama , Callegari Daihany Moraes , dos Reis Sávio Pinho , do Socorro Alves Namias Érika , da Cunha Ferreira Solange , de Souza Cláudia Regina Batista 
Published: April 2020

Background:
bZIP proteins participate in the regulation of gene expression, playing crucial
roles in various biological processes in plants, including response to environmental changes.
Luminosity is an environmental factor of extreme importance for plant metabolism, acting as a regulator
of its growth and development. Despite advances in the identification of bZIP proteins in several
plant species, studies on these transcription factors in cassava are lacking. Cassava (Manihot esculenta
Crantz) is one of the most important food crops in tropical and subtropical regions, mainly
in developing countries, where its storage root is a major source of calories for low-income people.

Objectives:
Our main aim was the isolation of a cDNA sequence encoding a bZIP protein from cassava
(MebZIP) as well as the in silico characterization of its nucleotide and deduced amino acid sequences.
In addition, we evaluated the expression pattern of the MebZIP gene in response to light,
and its possible relationship with regulation of the chalcone synthase (MeCHS) gene.

Method:
RT-PCR and 3’ and 5’ RACE assays were used to isolate the full-length cDNA sequence
of MebZIP. Bioinformatics tools were used to characterize the nucleotide and amino acid sequences
of MebZIP. Semiquantitative RT-PCR assays were used to evaluate the expression levels
of MebZIP and MeCHS genes.

Results:
We isolated the full-length cDNA sequence of MebZIP with a 1320-bp ORF encoding a
deduced protein with a predicted molecular weight and isoelectric point of 47 kDa and 5.85, respectively.
Comparative analyses with GenBank sequences showed high identity of MebZIP with bZIP
CPRF-2 of Hevea brasiliensis (XP_021650934) and Petroselinum crispum (Q99090.2). Besides
the basic region and leucine zipper domains, MebZIP contains putative conserved domains (D1-
D4), found in parsley CPRF-2 and bZIP proteins closely related to this protein. Since CPRF proteins
are known for their function in regulation of the CHS gene by light, we evaluated the expression
levels of the MebZIP gene and the possible target gene to be regulated by MebZIP (the
MeCHS gene) in cassava under light conditions. Semi-quantitative RT-PCR assays revealed that
MebZIP transcription increased in response to white light, with maximum expression levels at 6 h
of light exposure. On the other hand, the expression levels of the MeCHS gene were statistically
constant in all samples, indicating that they were not influenced by the experimental conditions
used here.

Conclusion:
The putative MebZIP protein identified in this work contains the conserved domains
(bZIP, D1-D4) that indicate its functionality, thus allowing it to be considered a new member of
the bZIP transcription factor CPRF-2 family. The expression levels of the MebZIP gene increased
during white light exposure, indicating a potential function in light-response in cassava.