Endogenous rhythmic growth in oak trees is regulated by internal clocks rather than resource availability

Abstract

Common oak trees display endogenous rhythmic growth with alternating shoot and root flushes. To explore the mechanisms involved, microcuttings of the Quercus robur L. clone DF159 were used for (13)C/(15)N labelling in combination with RNA sequencing (RNASeq) transcript profiling of shoots and roots. The effect of plant internal resource availability on the rhythmic growth of the cuttings was tested through inoculation with the ectomycorrhizal fungus Piloderma croceum. Shoot and root flushes were related to parallel shifts in above- and below-ground C and, to a lesser extent, N allocation. Increased plant internal resource availability by P. croceum inoculation with enhanced plant growth affected neither the rhythmic growth nor the associated resource allocation patterns. Two shifts in transcript abundance were identified during root and shoot growth cessation, and most concerned genes were down-regulated. Inoculation with P. croceum suppressed these transcript shifts in roots, but not in shoots. To identify core processes governing the rhythmic growth, functions [Gene Ontology (GO) terms] of the genes differentially expressed during the growth cessation in both leaves and roots of non-inoculated plants and leaves of P. croceum-inoculated plants were examined. Besides genes related to resource acquisition and cell development, which might reflect rather than trigger rhythmic growth, genes involved in signalling and/or regulated by the circadian clock were identified. The results indicate that rhythmic growth involves dramatic oscillations in plant metabolism and gene regulation between below- and above-ground parts. Ectomycorrhizal symbiosis may play a previously unsuspected role in smoothing these oscillations without modifying the rhythmic growth pattern.

Keywords: Ectomycorrhiza; Piloderma croceum; Quercus robur; RNASeq; growth cessation; stable isotope labelling..

Fig. 1.

Developmental stages of Quercus robur L. microcuttings. Developmental stages A (bud rest), B (bud swelling), C (shoot elongation), and D (leaf expansion) showing source leaves (SourceLeaf-1 and SourceLeaf-2) and sink leaves (SinkLeaf).

Fig. 2.

Biomass and both ¹³C and ¹⁵N excess of oak plant fractions at the four developmental stages. Measurements performed for harvested plant fractions [SinkLeaf (light green), SourceLeaf-1 (med green), and SourceLeaf-2 (dark green), stem (ochre), principal roots (dark brown), and lateral roots (light brown)] at the four developmental stages (A, B, C, and D): DWs in (A) control and (B) Piloderma croceum-inoculated oak microcuttings; ¹³C excess levels of (C) control and (D) P. croceum-inoculated plants; and ¹⁵N excess levels of (E) control and (F) P. croceum-inoculated plants. Root to shoot ratios (R/S_DW, R/S_13C, and R/S_15N) determined for plants at each of the four stages under each treatment are shown (means ±SE). Significant differences between stages are indicated with different letters for whole control plants (a, b), shoots (a’, b’), and roots (a’’, b’’) and, similarly, but in italics, for P. croceum-inoculated plants; Tukey test at P<0.05.

Fig. 3.

Venn diagrams based on up- and down-regulated contigs in leaves (Leaf) and lateral roots (LRs) within the Control (Cont) and P. croceum-inoculated (Pi) oak microcuttings. Differential expression of contigs was calculated in leaves (Leaf) by pairwise comparisons from stages D to A (_DtoA) at shoot growth cessation and in LRs from stages B to C (_BtoC) at root growth cessation. The blue ellipse marks 284 DECs in inoculated plants at leaf growth cessation, the yellow ellipse 1196 DECs in control plants at leaf growth cessation, and the red ellipse 1752 DECs of control plants during root growth cessation. Green colour indicates the ‘Leaf specific pool’ with DECs exclusively present at leaf growth cessation in control and inoculated plants, orange the ‘Cont specific pool’ with DECs exclusively present in control plants at shoot and root growth cessation, and beige the ‘Common pool’ with DECs belonging to control and inoculated plants at shoot and root growth cessation.

Fig. 4.

Profiles of transcript abundance of selected differentially expressed contigs (DECs) within ‘Common pool’, ‘Cont specific pool’, and ‘Leaf specific pool’ resulting from the intersections Cont&Pi_Leaf and Cont_Leaf&LR at growth cessation: in the ‘Common pool’, DECs are common to LRs and leaves of controls and leaves of inoculated oak microcuttings; in the ‘Leaf specific pool’ DECs are common to control and inoculated leaves; and in the ‘Cont specific pool’ DECs are common to leaves and LRs of control plants. The transcript abundance is given for leaves (Leaf) and roots (LR) of control (Cont) and P. croceum-inoculated (Pi) plants at the four developmental stages A (bud rest), B (bud swelling), C (shoot elongation), and D (leaf expansion). Except for ‘si’ (indicating SinkLeaves), all leaves were SourceLeaf-1. Significant differential expression of transcripts between stages at P<0.01 is given in bold lines.