Collaboration including OpenPlant researchers discovers that C4 photosynthesis has co-opted an ancient C3 regulatory code

C4Maize_Ninghui Shi_CC BY-SA 3.0.jpg

A new publication in Molecular Biology and Evolution has resulted from a collaboration of OpenPlant PI Prof. Julian Hibberd and researcher Dr Ivan Reyna-Llorens with colleagues in Portugal at the Instituto de Tecnologia Química e Biológica António Xavier, Universidade Nova de Lisboa, and the Instituto de Biologia Experimental e Tecnológica in Portugal. The paper shows that C4 photosynthesis has co-opted an ancient C3 regulatory code:

Borba AR, Serra TS, Górska A, Gouveia P, Cordeiro AM, Reyna-Llorens I, Kneřová J, Barros PM, Abreu IA, Oliveira MM, Hibberd JM, Saibo NJM (2018). Synergistic binding of bHLH transcription factors to the promoter of the maize NADP-ME gene used in C4 photosynthesis is based on an ancient code found in the ancestral C3 state. Molecular Biology and Evolution, msy060,


C4 photosynthesis has evolved repeatedly from the ancestral C3 state to generate a carbon concentrating mechanism that increases photosynthetic efficiency. This specialised form of photosynthesis is particularly common in the PACMAD clade of grasses, and is used by many of the world’s most productive crops. The C4 cycle is accomplished through cell-type specific accumulation of enzymes but cis-elements and transcription factors controlling C4 photosynthesis remain largely unknown. Using the NADP-Malic Enzyme (NADP-ME) gene as a model we tested whether mechanisms impacting on transcription in C4 plants evolved from ancestral components found in C3 species. Two basic Helix-Loop-Helix (bHLH) transcription factors, ZmbHLH128 and ZmbHLH129, were shown to bind the C4NADP-ME promoter from maize. These proteins form heterodimers and ZmbHLH129 impairs trans-activation by ZmbHLH128. Electrophoretic mobility shift assays indicate that a pair of cis-elements separated by a seven base pair spacer synergistically bind either ZmbHLH128 or ZmbHLH129. This pair of cis-elements is found in both C3 and C4 Panicoid grass species of the PACMAD clade. Our analysis is consistent with this cis-element pair originating from a single motif present in the ancestral C3 state. We conclude that C4 photosynthesis has co-opted an ancient C3 regulatory code built on G-box recognition by bHLH to regulate the NADP-ME gene. More broadly, our findings also contribute to the understanding of gene regulatory networks controlling C4 photosynthesis.

Image by Ninghui Shi: Cross section of a C4 plant. Specifically of a maize leaf. Vascular bundles shown. Drawing based on microscopic images courtesy of Cambridge University Plant Sciences Department. Image is shared under licence CC BY-SA 3.0