BIOLOGICAL OVERVIEW

Crocodile has recently joined Forkhead and Sloppy paired as identified forkhead domain proteins in Drosophila. Forkhead is involved in the specification of the posterior and anterior ectodermal gut primordia and regulates the invagination process that takes place during gut formation (Weigel, 1989). Sloppy paired 1 and Sloppy paired 2 appear to combine the functions of gap, pair rule and segment polarity classes of segmentation proteins. Slp functions as a gap-like gene in the prospective head region of the embryo and exerts segment polarity function in both the head and trunk anlage (Grossniklaus, 1994).

Crocodile has a dual role in Drosophila morphogenesis. An early role is in establishing the domains of expression for wingless and engrailed in the clypeolabrum, the anterior most segment of the fly's head. The clypeolabrum of croc mutant embryos is at least partially differentiated because the characteristic labral sensory organs; the labrum itself can be observed in croc mutant larvae. The role of croc early expression can be observed by examining internal clypeolabral structures. Two dorsal rows of muscle attachment sites, known as apodemes, are present in croc mutant embryos, while the single ventral row of apodemes is missing. In addition, the palisade-like structure of the dorsopharyngeal muscles never forms. Thus croc is required for the establishment of normal dorso-pharyngeal muscle pattern (Häcker, 1995).

Subsequent to its early role in determining muscle attachment sites, crocodile plays a role in the structuring of the intercalary segment. croc expression is maintained in intercalary derivatives, eventually seen in a row of cells lining the posterior wall of the pharynx. Larval cuticle preparations show that the posterior wall of the pharynx is absent in croc mutants, and the ventral arm of the cephalopharyngeal skeleton is strongly reduced (Häcker, 1995).

These two roles of croc appear to be independent of one another. The function of croc in the intercalary segment is unlikely to be due to Croc's role in regulating en and wg. Since the wg and en expression patterns are normal in the intercalary segment anlage of croc mutant embryos, Croc's role in that segment must be due to regulation of a different set of genes that are currently unknown. Its early activity is transiently required for determinative events in etablishing the posterior part of the clypeolabrum, as reflected in the altered segment polarity gene expression patterns of en and wg, while its activity in the developing intercalary segment is not required for the establishment of the segment anlage per se, but rather for the differentiation of specific structural elements (Häcker, 1995).

Croc is likely to play a role in development of the brain and central nervous system that has yet to be documented. croc is expressed in precursors of the ventral cord in a segmentally repeated fashion (Häcker, 1992).

GENE STRUCTURE

Bases in 5' UTR - 284

Exons - 1

Bases in 3' UTR - 628

PROTEIN STRUCTURE

Amino Acids - 529

Structural Domains

The forkhead domain, located in the N-terminal region of the Crocodile protein, shows a higher degree of sequence identity to the forkhead domains of FKH1 and MEH-1 of mouse (Kaestner, 1993 and Miura, 1993) and XFD-4 of Xenopus (Scheucher, 1996) as compared with the forkhead domains of either Forkhead itself or HNF-3alpha (Häcker, 1995).

A single-site mutation within the croc fkh domain, which causes a replacement of the first out of four conserved amino acid residues thought to be involved in the coordinate binding of Mg2+, abolishes the DNA binding of the protein in vitro. In view of the resulting lack-of-function mutant phenotype, it appears likely that metal binding by the affected region of the fkh domain is crucial for proper folding of the DNA-binding structure (Häcker, 1995).

The three-dimensional structure of an HNF-3/fork head DNA-recognition motif complexed with DNA has been determined by X-ray crystallography at 2.5 A resolution. This alpha/beta protein binds B-DNA as a monomer, through interactions with the DNA backbone and through both direct and water-mediated major and minor groove base contacts, inducing a 13 degrees bend. The transcription factor fold is very similar to the structure of histone H5. In its amino-terminal half, three alpha-helices adopt a compact structure that presents the third helix to the major groove. The remainder of the protein includes a twisted, antiparallel beta-structure and a random coil that interacts with the minor groove (Clark, 1993).

date revised:  25 April 2024   

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