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Ectopic shoot meristem generation in monocotyledonous rpk1 mutants is linked to SAM loss and altered seedling morphology.

Fiesselmann BS, Luichtl M, Yang X, Matthes M, Peis O, Torres-Ruiz RA - BMC Plant Biol. (2015)

Bottom Line: In strong rpk1 alleles, about 10 % of these (i. e. 1 % of all homozygotes) did not develop a SAM.The results highlight the developmental autonomy of the SAM vs. cotyledons and suggest that the primary rpk1 defect does not lie in the seedling's ability to express SAM genes or to develop a shoot meristem.The specific cotyledon defect in rpk1 mutants thus sheds light upon the developmental implications of the transition from two cotyledons to one.

View Article: PubMed Central - PubMed

Affiliation: Lehrstuhl für Genetik, Technische Universität München, Wissenschaftszentrum Weihenstephan, Emil-Ramann-Str. 8, D-85354, Freising, Germany. birgit.fiesselmann@gmail.com.

ABSTRACT

Background: In dicot Arabidopsis thaliana embryos two cotyledons develop largely autonomously from the shoot apical meristem (SAM). Recessive mutations in the Arabidopsis receptor-like kinase RPK1 lead to monocotyledonous seedlings, with low (10 %) penetrance due to complex functional redundancy. In strong rpk1 alleles, about 10 % of these (i. e. 1 % of all homozygotes) did not develop a SAM. We wondered whether RPK1 might also control SAM gene expression and SAM generation in addition to its known stochastic impact on cell division and PINFORMED1 (PIN1) polarity in the epidermis.

Results: SAM-less seedlings developed a simple morphology with a straight and continuous hypocotyl-cotyledon structure lacking a recognizable epicotyl. According to rpk1's auxin-related PIN1 defect, the seedlings displayed defects in the vascular tissue. Surprisingly, SAM-less seedlings variably expressed essential SAM specific genes along the hypocotyl-cotyledon structure up into the cotyledon lamina. Few were even capable of developing an ectopic shoot meristem (eSM) on top of the cotyledon.

Conclusions: The results highlight the developmental autonomy of the SAM vs. cotyledons and suggest that the primary rpk1 defect does not lie in the seedling's ability to express SAM genes or to develop a shoot meristem. Rather, rpk1's known defects in cell division and auxin homeostasis, by disturbed PIN1 polarity, impact on SAM and organ generation. In early embryo stages this failure generates a simplified monocotyledonous morphology. Once generated, this likely entails a loss of positional information that in turn affects the spatiotemporal development of the SAM. SAM-bearing and SAM-less monocotyledonous phenotypes show morphological similarities either to real monocots or to dicot species, which only develop one cotyledon. The specific cotyledon defect in rpk1 mutants thus sheds light upon the developmental implications of the transition from two cotyledons to one.

No MeSH data available.


Related in: MedlinePlus

RT-PCR analysis of monocot rpk1-7 seedlings with and without SAM. a Analysis of complete seedlings with (+ SAM) and without (− SAM) shoot meristem. RT-PCR amplification products after 40 cycles with primer pairs of genes as indicated. Note, that the expression of KNAT1 and 2 was present but very weak in seedlings with SAM. b Analysis of rpk1-7 monocot (− SAM) and rpk1-7 and wild-type dicot seedlings (+ SAM) separated into cotyledon tissue (Cot.) and (epi- and) hypocotyl and root tissue respectively (Rest)
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Fig4: RT-PCR analysis of monocot rpk1-7 seedlings with and without SAM. a Analysis of complete seedlings with (+ SAM) and without (− SAM) shoot meristem. RT-PCR amplification products after 40 cycles with primer pairs of genes as indicated. Note, that the expression of KNAT1 and 2 was present but very weak in seedlings with SAM. b Analysis of rpk1-7 monocot (− SAM) and rpk1-7 and wild-type dicot seedlings (+ SAM) separated into cotyledon tissue (Cot.) and (epi- and) hypocotyl and root tissue respectively (Rest)

Mentions: Next we analysed expression of SAM-specific genes such as WUS, STM, KNAT1 and KNAT2 (Fig. 4a) by semi-quantitative RT-PCR (see Methods). In this and other experiments care was taken that SAM-less seedlings were in fact devoid of a recognizable (late) SAM and that experiments with separated cotyledon tissue were not contaminated with hypocotyl and root tissue (see Methods). The cotyledon and leaf specific AS1 [27, 28] was included as control (in addition to ACT2). In one experiment, two seedlings of the SAM-less and two of the SAM-bearing group were separately analysed (including those shown in Fig. 1a to 1c). SAM-less seedlings expressed three of the four SAM-specific genes together with AS1, which was strongly expressed (Fig. 4a). While WUS was not found in these SAM-less seedlings, STM, KNAT1 and KNAT2 appeared to be aberrantly expressed in comparison to monocot seedlings with SAMs (Fig. 4a). The aliquots of both AS1 and ACT2 displayed significantly stronger expression since these genes have an overall expression in the cotyledon and the rest of the seedling respectively. Testing STM and AS1 (and AS2, not shown) in pools of cotyledons separated from the rest of the body, showed STM expression in cotyledons of SAM-less seedlings but not in those of controls (Fig. 4b). In addition, STM expression was also found in the rest of SAM-less monocots and as expected in the two controls (Fig. 4b). All bands had the expected sizes (as derived from the known transcripts). Additionally, representative bands were sequence verified. The expression of STM in both groups of monocot seedlings was comparable. A similar result was obtained using material of single seedlings (Additional file 1: Figure S2).Fig. 4


Ectopic shoot meristem generation in monocotyledonous rpk1 mutants is linked to SAM loss and altered seedling morphology.

Fiesselmann BS, Luichtl M, Yang X, Matthes M, Peis O, Torres-Ruiz RA - BMC Plant Biol. (2015)

RT-PCR analysis of monocot rpk1-7 seedlings with and without SAM. a Analysis of complete seedlings with (+ SAM) and without (− SAM) shoot meristem. RT-PCR amplification products after 40 cycles with primer pairs of genes as indicated. Note, that the expression of KNAT1 and 2 was present but very weak in seedlings with SAM. b Analysis of rpk1-7 monocot (− SAM) and rpk1-7 and wild-type dicot seedlings (+ SAM) separated into cotyledon tissue (Cot.) and (epi- and) hypocotyl and root tissue respectively (Rest)
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4492102&req=5

Fig4: RT-PCR analysis of monocot rpk1-7 seedlings with and without SAM. a Analysis of complete seedlings with (+ SAM) and without (− SAM) shoot meristem. RT-PCR amplification products after 40 cycles with primer pairs of genes as indicated. Note, that the expression of KNAT1 and 2 was present but very weak in seedlings with SAM. b Analysis of rpk1-7 monocot (− SAM) and rpk1-7 and wild-type dicot seedlings (+ SAM) separated into cotyledon tissue (Cot.) and (epi- and) hypocotyl and root tissue respectively (Rest)
Mentions: Next we analysed expression of SAM-specific genes such as WUS, STM, KNAT1 and KNAT2 (Fig. 4a) by semi-quantitative RT-PCR (see Methods). In this and other experiments care was taken that SAM-less seedlings were in fact devoid of a recognizable (late) SAM and that experiments with separated cotyledon tissue were not contaminated with hypocotyl and root tissue (see Methods). The cotyledon and leaf specific AS1 [27, 28] was included as control (in addition to ACT2). In one experiment, two seedlings of the SAM-less and two of the SAM-bearing group were separately analysed (including those shown in Fig. 1a to 1c). SAM-less seedlings expressed three of the four SAM-specific genes together with AS1, which was strongly expressed (Fig. 4a). While WUS was not found in these SAM-less seedlings, STM, KNAT1 and KNAT2 appeared to be aberrantly expressed in comparison to monocot seedlings with SAMs (Fig. 4a). The aliquots of both AS1 and ACT2 displayed significantly stronger expression since these genes have an overall expression in the cotyledon and the rest of the seedling respectively. Testing STM and AS1 (and AS2, not shown) in pools of cotyledons separated from the rest of the body, showed STM expression in cotyledons of SAM-less seedlings but not in those of controls (Fig. 4b). In addition, STM expression was also found in the rest of SAM-less monocots and as expected in the two controls (Fig. 4b). All bands had the expected sizes (as derived from the known transcripts). Additionally, representative bands were sequence verified. The expression of STM in both groups of monocot seedlings was comparable. A similar result was obtained using material of single seedlings (Additional file 1: Figure S2).Fig. 4

Bottom Line: In strong rpk1 alleles, about 10 % of these (i. e. 1 % of all homozygotes) did not develop a SAM.The results highlight the developmental autonomy of the SAM vs. cotyledons and suggest that the primary rpk1 defect does not lie in the seedling's ability to express SAM genes or to develop a shoot meristem.The specific cotyledon defect in rpk1 mutants thus sheds light upon the developmental implications of the transition from two cotyledons to one.

View Article: PubMed Central - PubMed

Affiliation: Lehrstuhl für Genetik, Technische Universität München, Wissenschaftszentrum Weihenstephan, Emil-Ramann-Str. 8, D-85354, Freising, Germany. birgit.fiesselmann@gmail.com.

ABSTRACT

Background: In dicot Arabidopsis thaliana embryos two cotyledons develop largely autonomously from the shoot apical meristem (SAM). Recessive mutations in the Arabidopsis receptor-like kinase RPK1 lead to monocotyledonous seedlings, with low (10 %) penetrance due to complex functional redundancy. In strong rpk1 alleles, about 10 % of these (i. e. 1 % of all homozygotes) did not develop a SAM. We wondered whether RPK1 might also control SAM gene expression and SAM generation in addition to its known stochastic impact on cell division and PINFORMED1 (PIN1) polarity in the epidermis.

Results: SAM-less seedlings developed a simple morphology with a straight and continuous hypocotyl-cotyledon structure lacking a recognizable epicotyl. According to rpk1's auxin-related PIN1 defect, the seedlings displayed defects in the vascular tissue. Surprisingly, SAM-less seedlings variably expressed essential SAM specific genes along the hypocotyl-cotyledon structure up into the cotyledon lamina. Few were even capable of developing an ectopic shoot meristem (eSM) on top of the cotyledon.

Conclusions: The results highlight the developmental autonomy of the SAM vs. cotyledons and suggest that the primary rpk1 defect does not lie in the seedling's ability to express SAM genes or to develop a shoot meristem. Rather, rpk1's known defects in cell division and auxin homeostasis, by disturbed PIN1 polarity, impact on SAM and organ generation. In early embryo stages this failure generates a simplified monocotyledonous morphology. Once generated, this likely entails a loss of positional information that in turn affects the spatiotemporal development of the SAM. SAM-bearing and SAM-less monocotyledonous phenotypes show morphological similarities either to real monocots or to dicot species, which only develop one cotyledon. The specific cotyledon defect in rpk1 mutants thus sheds light upon the developmental implications of the transition from two cotyledons to one.

No MeSH data available.


Related in: MedlinePlus