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Differential regulation of Knotted1-like genes during establishment of the shoot apical meristem in Norway spruce (Picea abies).

Larsson E, Sitbon F, von Arnold S - Plant Cell Rep. (2012)

Bottom Line: This was made both under normal embryo development and under conditions of reduced SAM formation by treatment with the polar auxin transport inhibitor NPA.Concomitantly with the formation of an embryonic SAM, the HBK2 and HBK4 genes displayed a significant up-regulation that was delayed by NPA treatment.Together, the results suggest that HBK2 and HBK4 exert similar functions related to the SAM differentiation and somatic embryo development in Norway spruce, while HBK1 and HBK3 have more general functions during embryo development.

View Article: PubMed Central - PubMed

Affiliation: Department of Plant Biology and Forest Genetics, Uppsala BioCenter, Swedish University of Agricultural Sciences, P. O. Box 7080, 75007 Uppsala, Sweden. emma.larsson@slu.se

ABSTRACT
Establishment of the shoot apical meristem (SAM) in Arabidopsis embryos requires the KNOXI transcription factor SHOOT MERISTEMLESS. In Norway spruce (Picea abies), four KNOXI family members (HBK1, HBK2, HBK3 and HBK4) have been identified, but a corresponding role in SAM development has not been demonstrated. As a first step to differentiate between the functions of the four Norway spruce HBK genes, we have here analyzed their expression profiles during the process of somatic embryo development. This was made both under normal embryo development and under conditions of reduced SAM formation by treatment with the polar auxin transport inhibitor NPA. Concomitantly with the formation of an embryonic SAM, the HBK2 and HBK4 genes displayed a significant up-regulation that was delayed by NPA treatment. In contrast, HBK1 and HBK3 were up-regulated prior to SAM formation, and their temporal expression was not affected by NPA. Ectopic expression of the four HBK genes in transgenic Arabidopsis plants further supported similar functions of HBK2 and HBK4, distinct from those of HBK1 and HBK3. Together, the results suggest that HBK2 and HBK4 exert similar functions related to the SAM differentiation and somatic embryo development in Norway spruce, while HBK1 and HBK3 have more general functions during embryo development.

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RT-PCR analysis of HBK1, HBK2, HBK3 and HBK4 expression in embryogenic cultures that were either competent to form fully mature cotyledonary embryos (28:05) or blocked at the PEM-to-embryo transition stage (88:1). Cell lines were sampled after 1 week of proliferation in the presence of PGRs (1), after 1 week in pre-maturation medium (2) and after 1 week on maturation medium (3). For cell line 28:05, these stages represent stages 1–3 in Fig. 1. The expression of PHOSPHOGLUCOMUTASE was used as a reference and H2O was used as a negative control in the RT-PCR analysis
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Fig3: RT-PCR analysis of HBK1, HBK2, HBK3 and HBK4 expression in embryogenic cultures that were either competent to form fully mature cotyledonary embryos (28:05) or blocked at the PEM-to-embryo transition stage (88:1). Cell lines were sampled after 1 week of proliferation in the presence of PGRs (1), after 1 week in pre-maturation medium (2) and after 1 week on maturation medium (3). For cell line 28:05, these stages represent stages 1–3 in Fig. 1. The expression of PHOSPHOGLUCOMUTASE was used as a reference and H2O was used as a negative control in the RT-PCR analysis

Mentions: It has been shown that HBK1 and HBK3 are expressed both in embryogenic cell lines that form fully mature cotyledonary embryos, and in blocked cell lines that never pass the PEM-to-embryo transition stage (Hjortswang et al. 2002). This is in contrast to HBK2, which is only expressed in embryogenic cell lines that form fully mature embryos. To further investigate the similarities and differences between the four HBK genes, the temporal expression of HBK4 was compared to that of HBK1, HBK2 and HBK3 in a control line (28:05) and in a blocked cell line (88:1). HBK1 and HBK3 were expressed in both cell lines, while HBK2 and HBK4 were expressed only in cell line 28:05 (Fig. 3). These results support a similar regulation of HBK1 and HBK3, and of HBK2 and HBK4. The results also reveal an intriguing, albeit tentative, link between the expression of HBK2 and HBK4 and proper embryo maturation, thus providing potential markers for somatic embryo competence.Fig. 3


Differential regulation of Knotted1-like genes during establishment of the shoot apical meristem in Norway spruce (Picea abies).

Larsson E, Sitbon F, von Arnold S - Plant Cell Rep. (2012)

RT-PCR analysis of HBK1, HBK2, HBK3 and HBK4 expression in embryogenic cultures that were either competent to form fully mature cotyledonary embryos (28:05) or blocked at the PEM-to-embryo transition stage (88:1). Cell lines were sampled after 1 week of proliferation in the presence of PGRs (1), after 1 week in pre-maturation medium (2) and after 1 week on maturation medium (3). For cell line 28:05, these stages represent stages 1–3 in Fig. 1. The expression of PHOSPHOGLUCOMUTASE was used as a reference and H2O was used as a negative control in the RT-PCR analysis
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3351593&req=5

Fig3: RT-PCR analysis of HBK1, HBK2, HBK3 and HBK4 expression in embryogenic cultures that were either competent to form fully mature cotyledonary embryos (28:05) or blocked at the PEM-to-embryo transition stage (88:1). Cell lines were sampled after 1 week of proliferation in the presence of PGRs (1), after 1 week in pre-maturation medium (2) and after 1 week on maturation medium (3). For cell line 28:05, these stages represent stages 1–3 in Fig. 1. The expression of PHOSPHOGLUCOMUTASE was used as a reference and H2O was used as a negative control in the RT-PCR analysis
Mentions: It has been shown that HBK1 and HBK3 are expressed both in embryogenic cell lines that form fully mature cotyledonary embryos, and in blocked cell lines that never pass the PEM-to-embryo transition stage (Hjortswang et al. 2002). This is in contrast to HBK2, which is only expressed in embryogenic cell lines that form fully mature embryos. To further investigate the similarities and differences between the four HBK genes, the temporal expression of HBK4 was compared to that of HBK1, HBK2 and HBK3 in a control line (28:05) and in a blocked cell line (88:1). HBK1 and HBK3 were expressed in both cell lines, while HBK2 and HBK4 were expressed only in cell line 28:05 (Fig. 3). These results support a similar regulation of HBK1 and HBK3, and of HBK2 and HBK4. The results also reveal an intriguing, albeit tentative, link between the expression of HBK2 and HBK4 and proper embryo maturation, thus providing potential markers for somatic embryo competence.Fig. 3

Bottom Line: This was made both under normal embryo development and under conditions of reduced SAM formation by treatment with the polar auxin transport inhibitor NPA.Concomitantly with the formation of an embryonic SAM, the HBK2 and HBK4 genes displayed a significant up-regulation that was delayed by NPA treatment.Together, the results suggest that HBK2 and HBK4 exert similar functions related to the SAM differentiation and somatic embryo development in Norway spruce, while HBK1 and HBK3 have more general functions during embryo development.

View Article: PubMed Central - PubMed

Affiliation: Department of Plant Biology and Forest Genetics, Uppsala BioCenter, Swedish University of Agricultural Sciences, P. O. Box 7080, 75007 Uppsala, Sweden. emma.larsson@slu.se

ABSTRACT
Establishment of the shoot apical meristem (SAM) in Arabidopsis embryos requires the KNOXI transcription factor SHOOT MERISTEMLESS. In Norway spruce (Picea abies), four KNOXI family members (HBK1, HBK2, HBK3 and HBK4) have been identified, but a corresponding role in SAM development has not been demonstrated. As a first step to differentiate between the functions of the four Norway spruce HBK genes, we have here analyzed their expression profiles during the process of somatic embryo development. This was made both under normal embryo development and under conditions of reduced SAM formation by treatment with the polar auxin transport inhibitor NPA. Concomitantly with the formation of an embryonic SAM, the HBK2 and HBK4 genes displayed a significant up-regulation that was delayed by NPA treatment. In contrast, HBK1 and HBK3 were up-regulated prior to SAM formation, and their temporal expression was not affected by NPA. Ectopic expression of the four HBK genes in transgenic Arabidopsis plants further supported similar functions of HBK2 and HBK4, distinct from those of HBK1 and HBK3. Together, the results suggest that HBK2 and HBK4 exert similar functions related to the SAM differentiation and somatic embryo development in Norway spruce, while HBK1 and HBK3 have more general functions during embryo development.

Show MeSH
Related in: MedlinePlus