Abstract
The Characeae are multicellular green algae with very close relationship to land plants. Their internodal cells have been the subject of numerous (electro-)physiological studies. When exposed to light, internodal cells display alternating bands of low and high pH along their surface in order to facilitate carbon uptake required for photosynthesis. Here we investigated for the first time the subcellular membrane protein composition of acidic and alkaline regions in internodal cells of Chara australis R. Br. using MS-proteomics. The identified peptides were annotated to Chara unigenes using a custom-made Chara database generated from a transcriptome analysis and to orthologous Arabidopsis genes using TAIR (The Arabidopsis Information Resource) database. Apart from providing the first public-available, functionally-annotated sequence database for Chara australis, the proteome study, which is supported by immunodetection, identified several membrane proteins associated with acidic regions that contain a high density of specific plasma membrane (PM) invaginations, the charasomes, which locally increase the membrane area to overcome diffusion limitation in membrane transport. An increased abundance of PM H+ ATPases at charasomes is consistent with their role in the acidification of the environment, but the characean PM H+ ATPase sequence suggests a different regulation compared to higher plant PM H+ ATPases. A higher abundance of H+ co-transporters in the charasome-rich, acidic regions possibly reflects enhanced uptake of ions and nutrients. The increase in mitochondrial proteins confirms earlier findings about the accumulation of cortical mitochondria in the acidic zones. The significant enrichment of clathrin heavy chains and clathrin adaptor proteins as well as other proteins involved in trafficking indicate a higher activity of membrane transport in the charasome-rich than in charasome-poor areas. New and unexpected data, for instance the upregulation and abundance of vacuolar transporters correlating with the charasome-rich, acidic cell regions account for new perspectives in the formation of charasomes.
Highlights
Since more than 6 decades, characean internodal cells have been used to study transport processes and to characterize transporter proteins by means of physiological techniques [1, 2], microscopy [3] and biochemistry including proteomics [4] and metabolomics [5]
Characean internodal cells produce extended acidic regions (H+ efflux) along their surface with alternating smaller alkaline areas with probably local OHefflux [2], required to achieve pH homoeostasis of the cytoplasm. This pH banding pattern can be visualized by phenol red [13] and acidic bands correlate with an enhanced photosynthesis compared to alkaline regions [14, 15] as well as high abundance and activity of cortical mitochondria [3] (S1 Fig)
A comprehensive view of the distribution of membrane and membrane-associated proteins between charasome-enriched regions of Chara cells helps to understand the physiological function of these specific plasma membrane regions in more detail
Summary
Since more than 6 decades, characean internodal cells have been used to study transport processes and to characterize transporter proteins by means of (electro-) physiological techniques [1, 2], microscopy [3] and biochemistry including proteomics [4] and metabolomics [5]. Characean internodal cells produce extended acidic regions (H+ efflux) along their surface with alternating smaller alkaline areas with probably local OHefflux [2], required to achieve pH homoeostasis of the cytoplasm. This pH banding pattern can be visualized by phenol red [13] and acidic bands correlate with an enhanced photosynthesis compared to alkaline regions [14, 15] as well as high abundance and activity of cortical mitochondria [3] (S1 Fig)
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