Analysis of the small chromosomal Prionium serratum (Cyperid) demonstrates the importance of reliable methods to differentiate between mono- and holocentricity

Abstract

For a long time, the Cyperid clade (Thurniceae-Juncaceae-Cyperaceae) was considered a group of species possessing holocentromeres exclusively. The basal phylogenetic position of Prionium serratum (Thunb.) Drège (Thurniceae) within Cyperids makes this species an important specimen to understand the centromere evolution within this clade. In contrast to the expectation, the chromosomal distribution of the centromere-specific histone H3 (CENH3), alpha-tubulin and different centromere-associated post-translational histone modifications (H3S10ph, H3S28ph and H2AT120ph) demonstrate a monocentromeric organisation of P. serratum chromosomes. Analysis of the high-copy repeat composition resulted in the identification of two centromere-localised satellite repeats. Hence, monocentricity was the ancestral condition for the Juncaceae-Cyperaceae-Thurniaceae Cyperid clade, and holocentricity in this clade has independently arisen at least twice after differentiation of the three families, once in Juncaceae and the other one in Cyperaceae. In this context, methods suitable for the identification of holocentromeres are discussed.

Keywords: CENH3/CENPA; Centromere type; Cyperids; Evolution; Holocentric chromosome; Thurniceae.

Fig. 1

Phylogenetic relationship of CENH3 between P. serratum and other plant species. The green and red branch represent monocot and eudicot species, respectively. The blue node indicates the reported holocentric species, and the sequences of the canonical histone H3 used as outgroup are shown in grey node. The CENH3 sequence accession numbers are listed in Suppl. Table 1

Fig. 2

Immunodetection of centromeric protein CENH3 (red) in P. serratum interphase nuclei (a), prophase (b) and its interaction with alpha-tubulin (green) in metaphase chromosomes (c, d). (d) Image taken by spatial structured illumination microscopy (SIM), enlargement (square) shows the interaction between CENH3 and alpha-tubulin

Fig. 3

Cell cycle–dependent, pericentromere-specific histone phosphorylated modification at H3S10 (a), H3S28 (b) and H2AT120 (c) in metaphase chromosomes of P. serratum. Overlapped signals between H3S10ph (green) and CENH3 (red) are shown in (a)

Fig. 4

Chromosome distribution of satellite DNA families and of CENH3 in P. serratum (2n = 46). Satellite repeat PsSat7 (a), PsSat41 (b), PsSat311 (c), PsSat157 (d), PsSat306 (e), Ps156a and Ps156b (f) were mapped on metaphase chromosomes. The fourth signal of PsSat41 is indicated by arrowheads (b). Colocalisations between PsSat306 and 45S rDNA and between Ps156a and Ps156b are shown in (e) and (f), respectively. The centromere specificity of Ps156a was confirmed by its overlapped signals, visualised in yellow in the merge images, with CENH3 in both interphase nuclei (g) and metaphase chromosomes (h). Image (g) was taken by structured illumination microscopy (SIM)

Fig. 5

Two types of DNA replication patterns in P. serratum shown by EdU labelling (red) and interphase nuclei counterstained with DAPI (blue). (a) Mainly uniform labelling and (b) clustered distribution of EdU signals