<em>NEEDLY</em>, a <em>Pinus radiata</em> ortholog of <em>FLORICAULA/LEAFY</em> genes, expressed in both reproductive and vegetative meristems
Abstract
The LEAFY/FLORICAULA genes from Arabidopsis and Antirrhinum are necessary for normal flower development and play a key role in diverse angiosperm species. A homologue of these flower meristem-identity genes, NEEDLY (NLY), has been identified in Pinus radiata. Although the NLY protein shares extensive sequence similarity with its angiosperm counterparts, it is lacking the proline-rich and acidic motifs thought to function as transcriptional activation domains. NLY already is expressed during vegetative development at least 5 years before the transition to the reproductive phase. Expression of NLY in transgenic Arabidopsis promotes floral fate, demonstrating that, despite its sequence divergence, NLY encodes a functional ortholog of the FLORICAULA/LEAFY genes of angiosperms. Expression of the LFY∷NLY transgene can largely complement the defects in flower development caused by a severe lfy allele.
Molecular and genetic studies have shown that the mechanisms controlling flower development largely are conserved across distantly related angiosperm plants (1). The first step in flower development is the switch from the vegetative phase, during which shoots and leaves are produced, to the reproductive phase, during which flowers are initiated. Once this switch has been made, flower meristem-identity genes promote the initiation of individual flowers. In Arabidopsis, these genes include LEAFY (LFY) (2), APETALA1 (AP1) (3), CAULIFLOWER (CAL) (4), APETALA2 (AP2) (5), and UNUSUAL FLORAL ORGANS (UFO) (6). At least two of these genes, LFY and AP1, not only are required for flower initiation but are also sufficient to induce flowering in lateral shoots when overexpressed in transgenic plants (7, 8).
The expression of LFY orthologs has been studied in detail in four angiosperm species. Of these, only expression of the snapdragon gene FLORICAULA (FLO) is specific to the reproductive phase (9), whereas the others are expressed, to varying degrees, during the vegetative phase as well. LFY expression during the vegetative phase is initially low but increases with the age of the plant. Expression levels are highest upon entering the reproductive phase, suggesting that LFY levels are critical for the transition to flowering. This point has been confirmed by demonstrating that increasing the copy number of endogenous LFY reduces the number of leaves produced before the first flower is formed (10). Both the Nicotiana NFL and the pea PEAFLO genes are expressed constitutively in emerging leaf primordia during the vegetative phase, as well as in floral organ primordia (11, 12). Although there is no evidence of a function of LFY and its ortholog in vegetative development of either Arabidopsis or Nicotiana, the situation in pea is different. Inactivation of PEAFLO in the pea mutant unifoliata (uni) not only causes a floral phenotype that is similar to that seen in flo or lfy mutants but also changes the morphology of the compound pea leaves (12).
Similar to many angiosperms, the “flowering” of P. radiata starts with the transformation of an indeterminate axillary apex into a determinate reproductive apex, which forms the strobili (cones) (13). A new long shoot terminal bud (LSTB) is formed at the tip of the rapidly elongating shoot during spring. The organogenic sequence of the apical meristem determines the fate of the shoot axis. The axillary apices that emerge on the sides of the apical meristem differentiate either as vegetative dwarf shoot buds (DSBs), reproductive pollen-cone buds (PCBs), or seed-cone buds (SCBs). SCBs become anatomically differentiated with the initiation of bracts (stage 1). Ovuliferous scale primordia are initiated from hypodermal cells on the adaxial base of bracts (stage 2). At stage 3, a fused bract-ovuliferous scale complex becomes displaced from the cone-bud axis. In differentiated PCBs, microsporangial initials appear in the peripheral zone of the axis (stage 1). During stage 2, microsporophyll initiation is complete and developed pollen mother cells are visible.
In contrast to angiosperms, our understanding of the molecular processes governing reproductive development in gymnosperms is very limited. A small family of MADS-box genes that is expressed in unisexual reproductive organs (and that shares similarity with floral organ-identity genes from angiosperms) has been isolated from two gymnosperms, Norway spruce (Picea abies) and Pinus radiata, and from their evolutionary ancestor, ferns (14–17). Here, we report the isolation and characterization of the first P. radiata gene belonging to the meristem-identity family of FLO/LFY-like genes. We show that the expression pattern of this gene, NEEDLY (NLY), is similar to that of these angiosperm genes, and we demonstrate with transgenic plants that it represents a true functional ortholog of the Arabidopsis LFY gene.
Acknowledgments
We thank Dr. Detlef Weigel (The Salk Institute) and Dr. Enrico Coen (John Innes Center) for kindly providing the LFY and FLO cDNAs; Dr. Detlef Weigel (The Salk Institute), Dr. Marty Yanofsky (University of California, San Diego), and Dr. Elena Alvarez-Buylla (University of Mexico) for very helpful discussions and for construction of the phylogenetic tree; Dr. Derek Harrison (University of Victoria) for many helpful comments; and Ms. Corinna Lange for assistance in preparation of manuscript.
ABBREVIATIONS
| SCB | seed-cone bud |
| PCB | pollen-cone bud |
| DSB | dwarf shoot bud |
| LSTB | long shoot terminal bud |
| LD | long day |
| SD | short day |
Footnotes
Data deposition: The sequence reported in this paper has been deposited in the GenBank database (accession no. {"type":"entrez-nucleotide","attrs":{"text":"U76757","term_id":"2160702"}}U76757).
References
- 1. Ma H. Genes Dev. 1994;8:745–756.[PubMed]
- 2. Weigel D, Alvarez J, Smyth D R, Yanofsky M F, Meyerowitz E M. Cell. 1992;69:843–859.[PubMed]
- 3. Mandel M A, Gustafson-Brown C, Savidge B, Yanofsky M E. Nature (London) 1992;360:273–277.[PubMed]
- 4. Kempin S A, Savidge B, Yanofsky M F. Science. 1995;267:522–525.[PubMed]
- 5. Jofuku K D, der Boer B G W, Van Montagu M, Okamuro J K. Plant Cell. 1994;6:1211–1225.
- 6. Ingram G C, Goodrich J, Wilkinson M D, Simon R, Haughn G W, Coen E S. Plant Cell. 1995;7:1501–1510.
- 7. Mandel M A, Yanofsky M F. Nature (London) 1995;377:522–524.[PubMed]
- 8. Weigel D, Nilsson O. Nature (London) 1995;377:495–500.[PubMed]
- 9. Bradley D, Vincent C, Carpenter R, Coen E. Development. 1996;122:1535–1544.[PubMed]
- 10. Blázquez M A, Soowal L N, Lee I, Weigel D. Development. 1997;124:3835–3844.[PubMed]
- 11. Kelly A J, Bonnlander M B, Meeks-Wagner D R. Plant Cell. 1995;7:225–234.
- 12. Hofer J, Turner L, Hellens R, Ambrose M, Matthews P, Michael A, Ellis N. Curr Biol. 1997;7:581–588.[PubMed]
- 13. Harrison D, Slee M. Can J For Res. 1992;22:1656–1668.[PubMed]
- 14. Tandre K, Albert V A, Sundas A, Engström P. Plant Mol Biol. 1995;27:69–87.[PubMed]
- 15. Mouradov A, Glassick T, Vivian-Smith A, Teasdale R D. Plant Physiol. 1996;110:1047.[PubMed]
- 16. Mouradov A, Glassick T, Teasdale R D. Plant Physiol. 1997;113:664.[PubMed]
- 17. Münster T, Pahnke J, Di Rosa A, Kim J T, Matin W, Saedler H, Theiβen G. Proc Natl Acad Sci USA. 1997;94:2415–2420.
- 18. Chang S, Puryear J, Cairney J. Plant Mol Biol Rep. 1993;11:113–116.[PubMed]
- 19. Jack T, Brockman L L, Meyerowitz E M. Cell. 1992;89:683–697.[PubMed]
- 20. Cardon G H, Frey M, Saedler H, Gierl A. Plant J. 1993;3:773–784.[PubMed]
- 21. Lee I, Wolfe D S, Nilsson O, Weigel D. Curr Biol. 1997;7:95–104.[PubMed]
- 22. Coen E, Romero J, Doyle S, Elliot R, Murphy G, Carpenter R. Cell. 1990;63:1311–1322.[PubMed]
- 23. Levin J Z, Meyerowitz E M. Plant Cell. 1995;7:529–548.