Kranz anatomical and biochemical characterization of C₄ photosynthesis in an aphyllous shrub Calligonum comosum (Polygonaceae)

Abstract

Calligonum is a shrubby and aphyllous C₄ eudicot lineage, employing green cylindrical shoots for carbon acquisition; however, comprehensive assessments of C₄ photosynthesis within the genus are limited. One species, C. comosum, grows in Jordan, offering an opportunity for evaluating features of C₄ photosynthesis in Calligonum. Light microscopy showed that C. comosum exhibits a variation of the classical salsoloid Kranz type (Calligonum variant). Transmission electron microscopy revealed traits typical of the NAD-malic enzyme (NAD-ME) subtype. Bundle sheath cells had numerous large mitochondria and mitochondria:chloroplast ratios were greater than those of mesophyll cells. Moreover, bundle sheath chloroplasts had developed grana, whereas mesophyll chloroplasts were deficient of grana. Enzyme activity and western blotting further supported NAD-ME as the principal decarboxylase. Immunolocalization of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco), phosphoenolpyruvate carboxylase (PEPC) and pyruvate-phosphate dikinase (PPdK) showed strict intercellular compartmentation patterns, with Rubisco confined to bundle sheath and PEPC and PPdK to mesophyll. Labeling for PEPC was observed in both the mesophyll and hypodermis, while PPdK was restricted to the mesophyll. We suggest classifying C. comosum as a NAD-ME subtype C₄ species and propose a mechanism of C₄ photosynthesis, suggesting a tractable C₄ model for further insights and studies on the biochemical intricacy of C₄ photosynthesis.

Keywords:

• Salsoloid Kranz anatomy
• enzyme activity
• immunolocalization
• granal development
• NAD-ME C₄ biochemical subtype
• Polygonaceae

Acknowledgements

The authors are grateful to Dr. A. El-Oqlah for help with the fieldwork, Fatima Al-Titi for help during the preparation of this manuscript, as well as the transmission electron microscope unit of Yarmouk University for use of the TEM facilities and for staff assistance.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Additional information

Funding

This research was funded by research grant number 10/2019 to R.M. from the Deanship of Scientific Research and Graduate Studies of Yarmouk University.