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The cloning and characterization of the CKNOX-A gene from Ceratopteris: The first isolated KNOTTED -like homeobox gene from a non-flowering plant
Homosporous ferns such as Ceratopteris richardii produce only one type of spore; irrespective of the genotype, spores can develop into either ameristic males, or meristic, hermaphroditic gametophytes. Determination of sexual identity occurs after spore germination by the action of the hormone antheridiogen, which promotes male development and is secreted by the hermaphrodite. Meristem development antagonizes the antheridiogen response, whereas activation of maleness by antheridiogen involves the specific repression of female-associated traits, such as the meristem and the archegonia.^ To elucidate the possible relationship between the meristem and the antheridiogen sex determination pathway, I cloned a Ceratopteris gene (CKNOX-A or CK) homologous to the homeobox-containing, meristem-specific KNOTTED1 or KN1 gene. CK encodes a putative protein that contains an 88-aa region 76% identical to the KN1 homeodomain. Expression studies performed by RT-PCR and in situ hybridization showed that CK is most strongly expressed in tissues that contain a meristem, such as hermaphroditic gametophytes, and sporophyte apices and leaves. However, CK RNA was also detected in antheridial cells in males. Surprisingly, the cellular localization of the CK hybridization signal differed depending on the regions of the CK gene used as a probe. Using the CK 3$\sp\prime$ UTR probe, the hybridization signal localized to the nuclei, in contrast to the cytoplasmic localization with a homeobox-containing probe.^ Functional homology of the CK gene to KN1 was assessed by overexpressing CK in Arabidopsis plants (ecotype Columbia). Transformant phenotypes ranged between two classes: in one, leaf size was severely reduced and plants formed petite rosettes. In the other, plants displayed a loss of apical dominance, by overproducing rosette leaves that developed without a visible phyllotactic pattern, or by forming several inflorescences simultaneously. Contrary to the KN overexpressing phenotypes in Arabidopsis, leaf shape was not altered by lobe formation in CK transformants. In conclusion, CK can be recognized by the molecules that control the initiation and maintenance of the Arabidopsis meristem, which suggests that Ceratopteris CK may play a similar role to the KNOTTED gene of maize. ^
Juarez, Cristina M, "The cloning and characterization of the CKNOX-A gene from Ceratopteris: The first isolated KNOTTED -like homeobox gene from a non-flowering plant" (1998). Doctoral Dissertations Available from Proquest. AAI9841882.