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  • 1.
    Hakman, Inger
    et al.
    University of Kalmar, School of Pure and Applied Natural Sciences.
    Hallberg, Henrik
    University of Kalmar, School of Pure and Applied Natural Sciences.
    Palovaara, Joakim
    University of Kalmar, School of Pure and Applied Natural Sciences.
    The polar auxin transport inhibitor NPA impairs embryo morphology and increases the expression of an auxin efflux facilitator protein PIN during Picea abies somatic embryo development2009In: Tree Physiology, ISSN 0829-318X, E-ISSN 1758-4469, Vol. 29, no 4, p. 483-496Article in journal (Refereed)
    Abstract [en]

    Auxin and polar auxin transport have been implicated in controlling embryo patterning and development in angiosperms but less is known from the gymnosperms. The aims of this study were to determine at what stages of conifer embryo development auxin and polar auxin transport are the most important for normal development and to analyze the changes in embryos after treatment with the polar auxin inhibitor N-1-naphthylphthalamic acid (NPA). For these studies, somatic embryos of Norway spruce (Picea abies L. Karst) were used. Growth on medium containing NPA leads to the formation of embryos with poor shoot apical meristem (SAM) and fused cotyledons, and to a pin-formed phenotype of the regenerated plantlets. The effect of NPA on embryo morphology was most severe if embryos were transferred to NPA-containing medium immediately before cotyledon initiation and SAM specification. Indole-3-acetic acid (IAA) was identified by immunolocalization in developing embryos. The highest staining intensity was seen in early staged embryos and then decreased as the embryos matured. No clear IAA-maxima was seen, although the apical parts of embryos, particularly the protoderm, and the suspensor cells appear to accumulate more IAA, as reflected by the staining pattern. The NPA treatment also caused expanded procambium and a broader root apical meristem in embryos, and a significant increase in the expression of a PIN1-like gene. Taken together, our results show that, for proper cotyledon initiation, correct auxin transport is needed only during a short period at the transition stage of embryo development, probably involving PIN efflux proteins and that a common mechanism is behind proper cotyledon formation within the species of angiosperms and conifers, despite their cotyledon number which normally differs.

  • 2.
    Hallberg, Henrik
    et al.
    University of Kalmar, School of Pure and Applied Natural Sciences.
    Hakman, Inger
    University of Kalmar, School of Pure and Applied Natural Sciences.
    Auxin localization and polar auxin transport during somatic embryogenesis in Norway spruce.2008Conference paper (Other academic)
  • 3.
    Hallberg, Henrik
    et al.
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Palovaara, Joakim
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Hakman, Inger
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Auxin polar transport and PIN localization pattern during conifer embryo development2010Conference paper (Other academic)
  • 4.
    Palovaara, Joakim
    et al.
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Hallberg, Henrik
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Stasolla, Claudio
    Plant Science, University of Manitoba.
    Hakman, Inger
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Comparative expression pattern analysis of WUSCHEL-related homeobox 2 (WOX2) and WOX8⁄9 in developing seeds and somatic embryos of the gymnosperm Picea abies2010In: New Phytologist, ISSN 0028-646X, E-ISSN 1469-8137, Vol. 188, no 1, p. 122-135Article in journal (Refereed)
    Abstract [en]

    In seed plants, current knowledge concerning embryonic pattern formation by polar auxin transport (PAT) and WUSCHEL-related homeobox (WOX) gene activity is primarily derived from studies on angiosperms, while less is known about these processes in gymnosperms. In view of the differences in their embryogeny, and the fact that somatic embryogenesis is used for mass propagation of conifers, a better understanding of embryo development is vital.

    The expression patterns of PaWOX2 and PaWOX8/9 were followed with quantitative reverse transcription–polymerase chain reaction (qRT-PCR) and in situ hybridization (ISH) during seed and somatic embryo development in Norway spruce (Picea abies), and in somatic embryos treated with the PAT inhibitor N-1-naphthylphthalamic acid (NPA).

    Both PaWOX2 and PaWOX8/9 were highly expressed at the early growth stages of zygotic and somatic embryos, and shared a similar expression pattern over the entire embryo. At later embryo stages, high expression of PaWOX8/9 became restricted to cotyledon primordia, epidermis, procambium and root apical meristem (RAM), which became most evident in NPA-treated somatic embryos, while expression of PaWOX2 was much lower.

    Our results suggest an ancestral role of WOX in seed plant embryo development, and strengthen the proposed connection between PAT, PIN-FORMED (PIN) and WOX in the regulation of embryo patterning in seed plants.

     

     

     

     

     

     

     

     

     

     

     

     

     

     

     

     

     

     

     

     

     

     

     

     

     

     

     

     

     

  • 5.
    Palovaara, Joakim
    et al.
    University of Kalmar, School of Pure and Applied Natural Sciences.
    Hallberg, Henrik
    University of Kalmar, School of Pure and Applied Natural Sciences.
    Stasolla, Claudio
    Luit, Bert
    Hakman, Inger
    University of Kalmar, School of Pure and Applied Natural Sciences.
    Expression dynamics of PIN1 and WOX genes during Norway spruce (Picea abies) somatic embryogenesis.2009Conference paper (Other academic)
  • 6.
    Palovaara, Joakim
    et al.
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Hallberg, Henrik
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Stasolla, Claudio
    Plant Science, University of Manitoba.
    Luit, Bert
    Plant Science, University of Manitoba.
    Hakman, Inger
    Linnaeus University, Faculty of Science and Engineering, School of Natural Sciences.
    Expression of a gymnosperm PIN homologous gene correlates with auxin immunolocalization pattern at cotyledon formation and in demarcation of the procambium during Picea abies somatic embryo development and in seedling tissues2010In: Tree Physiology, ISSN 0829-318X, E-ISSN 1758-4469, Vol. 30, no 4, p. 479-489Article in journal (Refereed)
    Abstract [en]

    In seed plants, the body organization is established during embryogenesis and is uniform across gymnosperms and angiosperms, despite differences during early embryogeny. Evidence from angiosperms implicates the plant hormone auxin and its polar transport, mainly established by the PIN family of auxin efflux transporters, in the patterning of embryos. Here, PaPIN1 from Norway spruce (Picea abies [L.] Karst.), a gene widely expressed in conifer tissues and organs, was characterized and its expression and localization patterns were determined with reverse transcription polymerase chain reaction and in situ hybridization during somatic embryo development and in seedlings. PaPIN1 shares the predicted structure of other PIN proteins, but its central hydrophilic loop is longer than most PINs. In phylogenetic analyses, PaPIN1 clusters with Arabidopsis thaliana (L.) Heynh. PIN3, PIN4 and PIN7, but its expression pattern also suggests similarity to PIN1. The PaPIN1 expression signal was high in the protoderm of pre-cotyledonary embryos, but not if embryos were pre-treated with the auxin transport inhibitor N-1-naphthylphthalamic acid (NPA). This, together with a high auxin immunolocalization signal in this cell layer, suggests a role of PaPIN1 during cotyledon formation. At later stages, high PaPIN1 expression was observed in differentiating procambium, running from the tip of incipient cotyledons down through the embryo axis and to the root apical meristem (RAM), although the mode of RAM specification in conifer embryos differs from that of most angiosperms. Also, the PaPIN1 in situ signal was high in seedling root tips including root cap columella cells. The results thus suggest that PaPIN1 provides an ancient function associated with auxin transport and embryo pattern formation prior to the separation of angiosperms and gymnosperms, in spite of some morphological differences.

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