Original Article Role of Jun N-terminal Kinase (JNK) signaling in the wound healing and regeneration of a Drosophila melanogaster wing imaginal disc JAAKKO MATTILA* ,1 , LEONID OMELYANCHUK 2 , SATU KYTTÄLÄ 1 , HEIKKI TURUNEN 1 and SEPPO NOKKALA 1 1 Laboratory of Genetics, Department of Biology, University of Turku, Finland and 2 Laboratory of Cell Cycle Genetics, Institute of Cytology and Genetics, Novosibirsk, Russia ABSTRACT When a fragment of a Drosophila imaginal disc is cultured in growth permissive conditions, it either regenerates the missing structures or duplicates the pattern present in the fragment. This kind of pattern regulation is known to be epimorphic, i.e. the new pattern is generated by proliferation in a specialized tissue called the blastema. Pattern regulation is accompanied by the healing of the cut surfaces restoring the continuous epithelia. Wound healing has been considered to be the inductive signal to commence regenerative cell divisions. Although the general outlines of the proliferation dynamics in a regenerating imaginal disc blastema have been well studied, little is known about the mechanisms driving cells into the regenerative cell cycles. In this study, we have investigated the role of Jun N-terminal Kinase (JNK) signaling in the wound healing and regeneration of a Drosophila wing imaginal disc. By utilizing in vivo and in vitro culturing of incised and fragmented discs, we have been able to visualize the dynamics in cellular architecture and gene expression involved in the healing and regeneration process. Our results directly show that homotypic wound healing is not a prerequisite for regenerative cell divisions. We also show that JNK signaling participates in imaginal disc wound healing and is regulated by the physical dynamics of the process, as well as in recruiting cells into the regenerative cell cycles. A model describing the determination of blastema size is discussed. KEY WORDS: wing imaginal disc, regeneration, wound healing, Jun N-terminal Kinase (JNK) Introduction The imaginal discs of Drosophila melanogaster have an intriguing capability to regenerate missing structures. When cut into two pieces and cultured in the abdomens of adult flies, in general the larger piece undergoes regeneration while the comple- mentary piece duplicates (Bryant 1971, Schubiger 1971). An absolute requirement for pattern regulation (regeneration/dupli- cation) to occur is growth (Schubiger 1973). In particular, pattern regulation of an imaginal disc is known to be epimorphic (Dale & Bownes 1980, Bryant & Fraser 1988) (although suggestions to the contrary exist, see Adler 1981), i.e. the emerging of the new pattern is dependent on cell divisions taking place in a specialized proliferating tissue called the blastema. The dynamics of regen- erative proliferation have been extensively studied by clonal analysis (Abbot et al. 1981), incorporation of 3 H-thymidine into replicating cells (Dale & Bownes 1980, 1981, Adler 1984, Kiehle & Schubiger 1985, O’Brochta & Bryant 1987), direct cell counting Int. J. Dev. Biol. 49: 391-399 (2005) doi: 10.1387/ijdb.052006jm *Address correspondence to: Jaakko Mattila. Institute of Biotechnology, Viikinkaari 9 (P.O. Pox 56), 00014 University of Helsinki, Finland. Fax: +358-9-19159366. e-mail: jaakko.mattila@helsinki.fi Abbreviations used in this paper: GFP, green fluorescent protein; JNK, Jun N- terminal Kinase. 0214-6282/2005/$25.00 © UBC Press Printed in Spain www.intjdevbiol.com (Dunne 1981), volume measurement of cultured disc fragments (Adler 1981), flow cytometric analysis (Fain & Alvarez 1987) and immunolocalization of incorporated bromodeoxyuridine (Bryant & Fraser 1988). Regeneration of an imaginal disc is accompanied by the healing of the cut edge surfaces, first by transient heterotypic contacts of the peripodial membrane and columnar epithelia and later by homotypic healing as the cell layers fuse to restore the continuous epithelia (Reinhardt et al. 1977, Reinhardt & Bryant 1981). The prevailing view of pattern regulation is that apposition of differential positional values by homotypic healing induces intercalary growth resulting in either regeneration or duplication depending on the developmental capacity of the apposed cells. This idea, largely based on the polar co-ordinate model devel- oped by French et al. (1976), is however impaired by the finding