The anatomy of walnut staminate inflorescences from the time of initiation until budbreak has been studied by several investigators (Li et al., 2011; Luza and Polito, 1988). The consensus among these studies is that the development of a walnut staminate inflorescence requires ≈1 year from the time of initiation to maturation. However, the differentiation course varies by cultivar and climate (Polito and Katherine, 1997; Zhang et al., 1995). Marker characteristics should be linked genetically to the process of differentiation to ensure stability in the face of climatic variation. Attempting to project a marker event may be required. Markers that are easily duplicated and measured can be used to track events during histogenesis and enable comparisons of different studies. Morphological features that can be used as indicators of developmental stage have been identified in various plants. For example, flower bud development in Cyclocarya paliurus can be defined based on floret size, anther color, and characteristics of the stigma (Fu et al., 2011), whereas the emergence of a globular embryo is a reliable indicator of the beginning of tissue differentiation in pineapple [Ananas comosus (He et al., 2012)]. Evaluating development based on morphology is easy, but the relationship between the morphology and physiology of walnut catkins has not been established.
Auxin is an essential hormone that has been implicated in many aspects of plant growth and development (Woodward and Barrel, 2005), and the relationship between auxin and flower formation has been extensively studied (Aloni et al., 2006; Cheng and Zhao, 2007; Jiménez, 2005; Marleen and Eva, 2012). Auxin is considered to be a key regulator of floral development, but the exact role of auxin in this process has only recently come to be well understood (Chandler, 2011). Valentina et al. (2008) demonstrated that auxin synthesized in anthers coordinates anther dehiscence and pollen maturation, whereas auxin transport contributes to the independent regulation of preanthesis filament elongation; hence, auxin is an important physiological regulator of staminate flower differentiation. Using immunologic techniques, auxin can be detected in situ in plant tissues. Indeed, immunohistochemical localization studies have been performed in maize [Zea mays (Vysotskaya et al., 2007)], arabidopsis [Arabidopsis thaliana (Aloni et al., 2003)], the shoot apices of strawberry [Fragaria ×ananassa (Hou and Huang, 2005)], and hybrid poplar [Populus alba × (P. davidiana × P. simonii) × P. tomentosa (Dong et al., 2012)].
Previous attempts have been made to establish the relationships among the morphological characteristics, physiology, and histogenesis of staminate flowers in walnut (Vladimra and Sladky, 1971; Yates and Sparks, 1992). The application of exogenous hormones could transform the staminate primordia into vegetative buds. These findings comprise a first step toward elucidating the mechanism of walnut staminate flowering and the role of hormones in walnut catkin formation. Additionally, they have created a foundation for studying the regulation of staminate flowering in walnut. The results of this study also have significance for dioecious species, in which male and female individuals produce unisexual flowers and are characterized by sexual dimorphism (Cristiana Moliterni et al., 2004).
In this present study, an anti-indole acetic acid (IAA) monoclonal antibody was used to detect and localize endogenous IAA during walnut flower differentiation using an immunochemical approach. We provide a substantial base for the further research on the mechanism of IAA action during staminate flower bud differentiation.
Aloni, R.K., Schwalm, M. & Langhans, C.I. 2003 Gradual shifts in sites of free-auxin production during leaf-primordium development and their role in vascular differentiation and leaf morphogenesis in arabidopsis Planta 216 841 853
Cristiana Moliterni, V.M., Cattivelli, L., Ranalli, P. & Mandolino, G. 2004 The sexual differentiation of Cannabis sativa L.: A morphological and molecular study Euphytica 140 95 106
Dong, N.G., Yin, W.L., Gao, Y. & Pei, D. 2012 Indole-3-acetic acid accumulation during poplar rhizogenesis revealed by immunohistochemistry Biol. Plant. 56 581 584
Fu, X.X., Feng, L., Shang, X.L., Yang, W.X. & Fang, S.Z. 2011 Observation of morphological and anatomical characters on staminate and pistillate flower differentiation in Cyclocarya paliuru J. Nanjing For. Univ. (Natural Sci. Ed.) 35 17 22
Gregorio, O.A., Sonia, V.S., Aldebaran, C., Joseph, G.D. & Felipe, C.G. 2012 Inception of maleness: Auxin contribution to flower masculinization in the dioecious cactus Opuntia stenopetala Planta 236 225 238
He, Y.H., Fang, S.Q., Hu, Z.Y., Ma, J., Luo, J., Wu, C.H., Cao, L., Lu, M. & Chen, C.J. 2012 Morphological and anatomical analysis of pineapple somatic embryogenesis Acta Hort. Sinica 39 57 63
Heisler, M.G., Ohno, C., Das, P., Sieber, P., Reddy, G.V., Long, J.A. & Meyerowitz, E.M. 2005 Patterns of auxin transport and gene expression during primordium development revealed by live imaging of the arabidopsis Inflorescence meristem Curr. Biol. 15 1899 1911
Holgate, C.S., Jackson, P. & Cowen, P.N. 1983 Immunogold silver staining: New method of immunostaining with enhanced sensitivity J. Histochem. Cytochem. 7 938 944
Hou, Z.X. & Huang, W.D. 2005 Immunohistochemical localization of IAA and ABP1 in strawberry shoot apexes during floral induction Planta 222 678 687
Jiménez, V.M. 2005 Involvement of plant hormones and plant growth regulators on in vitro somatic embryogenesis Plant Growth Regulat. 47 91 110
Leverone, L.A., Stroup, T.L. & Caruso, J.L. 1991 Western blot analysis of cereal grain prolamins using an antibody to carboxyl-linked indoleacetic acid Plant Physiol. 96 1076 1078
Li, Y.T., Zhao, Y.G., Yang, K.Q., Fang, Y.Y. & Hou, L.Q. 2011 Studies on floral organ development process of precocious walnut (juglans regia) by anatomia Acta Hort. Sinica 38 434 440
Luza, J.G. & Polito, V.S. 1988 Microsporogenesis and anther differentiation in Juglans regia L.: A developmental basis for heterodichogamy in walnut Bot. Gaz. 149 30 36
Oka, M., Miyamoto, K., Okada, K. & Ueda, J. 1999 Auxin polar transport and flower formation in arabidopsis thaliana transformed with indoleacetamide hydrolase (IAAH) gene Plant Cell Physiol. 40 231 237
Orna, A.K., Cheng, J.C., Chen, L.J., Moctezuma, E. & Sung, Z.R. 2002 Indole acetic acid distribution coincides with vascular differentiation pattern during arabidopsis leaf ontogeny Plant Physiol. 130 199 209
Polito, V.S. & Katherine, P. 1997 The relationship between phenology of pistillate flower organogenesis and mode of heterodichogamy in Juglans regia L. (Juglandaceae) Sex. Plant Reprod. 10 36 39
Valentina, C., Maria, M.A., Giuseppina, F., Paolo, C. & Maura, C. 2008 Auxin regulates arabidopsis anther dehiscence, pollen maturation, and filament elongation Plant Cell 20 1760 1774
Vladimra, L. & Sladky, Z. 1971 The role of growth regulators in the differentiation of walnut buds (Juglans regia L.) Biol. Plant. 13 361 367
Vysotskaya, L.B., Veselov, S.Y., Veselov, D.S., Filippenko, V.N., Ivanov, E.A., Ivanov, I.I. & Kudoyarova, G.R. 2007 Immunohistological localization and quantification of IAA in studies of root growth regulation Russ. J. Plant Physiol. 54 827 832