Razumovskaya A.V. Cytology of the minor-vein phloem in 320 species from the subclass Asteridae suggests a high diversity of phloem-loading modes. Frontiers in Plant Science. 2013, V. 4, Article 312.

Batashev et al Minor vein phloem in Asteridae phloem initials in course of minor vein formation are periclinal; only the last divisions in the group of phloem cells are anticlinal (original micrographs shown in Gamalei, 1989, for Senecio sp.). As a result, the phloem cells form several layers (“tiers”) which are situated one under another (Figure 2B). These tiers consist either of SE-CCCs or of phloem parenchyma cells. Here, xylem and phloem are not separated by a layer of parenchyma cells. In the present study, the layout of minor veins in most stud­ ied species corresponded to either type 1 or 2 (Supplemental Table 1). The following traits were attributed to type 1: (i) minor veins show a well-defined spatial organization; (ii) the veins can contain companion cells of more than one structural type; (iii) the number of companion cells equals the number of sieve ele­ ments (CC:SE = 1) which results from the pattern of cell divisions during m inor vein formation; (iv) phloem initials do not form phloem parenchyma cells (but vascular parenchyma is present, see above) in the course of minor vein development. The minor veins of type 2 showed the following traits: (i) minor veins show a well-defined spatial organization, (ii) companion cells within the vein belong to a single structural type; (iii) the CC:SE ratio equals 2 or 1.5 (for instance when two SE-CCCs are combined in one minor vein, one with the ratio of CC:SE = 1 and the second with a ratio of CC:SE = 2); (iv) phloem parenchyma is present, and phloem parenchyma and SE-CCCs are positioned in alternating layers. Apart from these types, a small group of species with minor veins of variable composition resulting from unordered anticli­ nal and periclinal divisions during vein formation was found in the Asteridae. In this review, we refer to this group as type 0; it consists mostly of representatives of the l-2a group according to Gamalei (1989, 1991). In this type, (i) the spatial organization of m inor veins is not well-defined; (ii) the number of CCs exceeds the number of SEs; (iii) the CC:SE ratio is not stable; (iv) phloem parenchyma cells can be present or absent. Interestingly, compan­ ion cells in type 0 minor veins studied thus far were never found to contain highly specialized structures like plasmodesmal fields or a cell wall labyrinth. A published example of this minor vein organization is Digitalis grandiflora (Turgeon et al., 1993). In types 1 and 2, several subtypes can be distinguished on the basis of (i) presence of plasmodesmal fields at companion cell/bundle sheath boundary, (ii) type of plastids in companion cells, (iii) presence of cell wall ingrowths in companion cells, (iv) presence and position of phloem parenchyma cells within minor veins, and (v) presence of cell wall ingrowths in phloem parenchyma cells (Table 2). The presence or absence of these characteristics can be clearly seen on almost any transversal sec­ tion. This categorization does not include subordinate features such as type of plasmodesmata branching, type of transported sugars or presence of starch in leucoplasts of IC-like cells. The full information on the structural features of the minor vein phloem in the 320 analyzed species is shown in the Supplemental Table 1. Type 1 was divided into subtypes I-IV (Figures 3, 4A-D). In the subtype 1-1, the minor veins contain three SE-CCCs with companion cells of a similar structure (IC, ICL, or CC- a) (Table 1; Figure4A). This is one of the most widespread subtypes in the Asteridae occurring in the Acanthaceae, Apocynaceae, Bignoniaceae, Cornaceae, Eucommiaceae, Griseliniaceae, Gesneriaceae, Hydrangeaceae, Lamiaceae, Oleaceae, Orobanchaceae, Pawlowniaceae, Plantaginaceae, Rubiaceae, Scrophulariaceae, and Verbenaceae. Subtype 1-II is very close to 1-1; the only difference is that in minor veins with three SE-CCCs, the abaxial SE-CCC contains an OC-a, a companion cell without plasmodesmal fields and cell wall ingrowths (Figure4B). The subtype 1-II is also quite widespread and was found in Acanthaceae, Bignoniaceae, Lamiaceae, Orobanchaceae, Phrymaceae, Plantaginaceae, Rubiaceae, Table 2 | Classification of the minor vein phloem according to its spatial organization in leaves of Asteridae. Type Subtype Vein symmetry CC subtype Number of CC subtypes combined in one phloem ending Phloem parenchyma CWI in phloem parenchyma Examples 1 1-1 + IC, ICL, CC-a 1 - - Figure 4A 1-11 + IC,OC-a 2 - - Figure 4B 1-111 + (rarely -) IC or MIC-a TC-b 2 - - Figure 4C 1-IV + CC-b 1 - - Figure 4D 0 0 - ОС-a, OC-b 1 - (rarely +) - Figure 4F 2 2-1 + ОС-a, OC-b 1 + - Figure 5A 2-II + (rarely -) ОС-a, OC-b 1 + + Figure 5B 2-III + TC-a 1 + - Figure 5C 2-IV + TC-a 1 + + Figure 5D 2-V + TC-c 1 + - Figure 5E 2-VI +/- MIC-b 1 + - Figure 5F CWI, cell wall ingrowths; CC, companion cells. "+ " marks the presence and " the absence of a feature. www.frontiersin.org August 2013 | Volume 4 | Article 312 | 7