**1. Introduction**

Red seaweed cell-walls are mainly made of complex sulphated galactans primarily agar and carrageenans, which comprise a large family of hydrocolloids, depending on the varying degree of sulphation [1–3]. Specifically, carrageenans can be classified according to their gel-forming ability [4]. The differences among the various forms of carrageenan (<sup>κ</sup>-, ι- and λ-carrageenans) are related to the amount of sulphate groups, affecting solubility in water and the strength of carrageenans [5]. Moreover, cell walls have to change their carrageenan composition to adapt to the different development stages of the seaweed [6]. Thus, carragenophytic-alga gametophytes are known to be composed of κ- and ι carrageenan, whereas tetrasporophytes have λ-carrageenan [4].

Seaweed responses to the presence of reproductive structures is to push thallus cells to the sides, generate anatomical changes to locate reproductive structures and trigger fluctuations in the composition of the cell wall [4,7]. As the integrity of the cell wall is essential for maintaining growth and development of alga thalli and for the survival of reproductive structures, alterations in carrageenan composition can be assumed, as galactose units work together to build, incorporate and remove

sulphated groups to and from the growing strands of galactans. Furthermore, these changes in the amount of sulphate groups influence the ability of seaweed to model responses, such as the softening and flexibility of thalli and facilitating the growth in size of reproductive structures. Seaweed can also display responses for recovering of thallus status after sporogenesis.

Additionally, sporogenesis is inherently linked to stressors such as volatile growth regulators, tide periods, hours of irradiation and temperature [8–10]. These environmental stressors also have an impact on generating reactive oxygen species (ROS in the form of O2, H2O2 or OH−) [9,10]. Moreover, ROSs also play an important role in softening thalli and therefore also facilitate the development of cystocarps in red seaweed [8]. Reactive oxygen species act as signalling molecules under stress and induce cell responses [11,12]. Moreover proteins, such as WD40 and cytochrome P450, can be synthesised to reduce such oxidative damage [9,10,13]. Carrageenans, as components of the cell wall in seaweeds, have been reported to also show antioxidant activity by scavenging hydrogen peroxide [14,15].

Consistent with the chemical complexity of the cell wall and multiple environment stressors, the transcriptional machinery underlying synthesis and modification of cell-wall polysaccharides is intricate. Particularly in the case of carrageenans, the biosynthesis pathway has not been fully described, although three main classes of enzymes have been proposed namely galactosyltransferases, sulfotransferases and galactose 6 sulfurylases [16]. Significantly, carbohydrate sulfotransferase, which add a sulphate group from a donor molecule (often <sup>3</sup>phosphoadenosine-5phosphosulfate, PAPS), have been described in carrageenophytes. Galactose-6 sulfurylase, which catalyses the formation of the 3,6-anhydrogalactose residues by removing C6 sulphate group in sulphated galactans [17], has also been reported specifically in red macroalgae (Figure 1).

**Figure 1.** Schematic biosynthetic pathway for sulphate assimilation and synthesis of carrageenan. PAPS, <sup>3</sup>phosphoadenosine-5phosphosulfate.

*Grateloupia imbricata* is a carragenophytic red seaweed, with triphasic life cycle (Figure 2). This macroalga represents a candidate model organism for basic studies of physiology, owing to its ability to produce raw material such as carrageenan.

**Figure 2.** Diagram of a tri-genetic life cycle in the red alga *Grateloupia imbricata* comprising gametophytes (haploids), the carposporophyte, that develops on the female gametophyte after fertilisation, and the sporophyte (diploid). Taken from Garcia-Jimenez and Robaina (2019) DOI: http://dx.doi.org/10.5772/ intechopen.83353.

Transcriptome information of *G. imbricata* revealed the presence of transcripts required for biosynthesis of sulphated polysaccharides, so the study of these transcripts can contribute to the understanding of carrageenan synthesis [18]. This paper focuses on the characterisation of expression of genes such as *carbohydrate sulfotransferase* and *galactose 6 sulfurylase* that are specifically related to addition and removal of sulphate groups to galactan. We hypothesised that the expression of genes encoding enzymes of carrageenan synthesis in *G. imbricata* can be a starting point for further studies on sulphation of carrageenan. Moreover, these genes can be correlated to reproductive stage (carposporogenesis) of *G. imbricata*. Our aim is to show that the expression of two genes involved in the sulphation (*carbohydrate sulfotransferase*) and desulphation (*galactose-6 sulfurylase*) of the galactan backbone is related to the stage of development of thalli and to reproductive structures (cystocarps) through flexibility and softening of thalli. Likewise, we determine to what extent genes encoding stress proteins (*Cytochrome P450* and *WD 40*) can also be involved during cystocarp maturation, as the presence of cystocarps would also comprise the cell wall softening.
