Distinct functions are implicated for the GATA-4, -5, and -6 transcription factors in the regulation of intestine epithelial cell differentiation.

Based on conserved expression patterns, three members of the GATA family of transcriptional regulatory proteins, GATA-4, -5, and -6, are thought to be involved in the regulation of cardiogenesis and gut development. Functions for these factors are known in the heart, but relatively little is understood regarding their possible roles in the regulation of gut-specific gene expression. In this study, we analyze the expression and function of GATA-4, -5, and -6 using three separate but complementary vertebrate systems, and the results support a function for these proteins in regulating the terminal-differentiation program of intestinal epithelial cells. We show that xGATA-4, -5, and -6 can stimulate directly activity of the promoter for the intestinal fatty acid-binding protein (xIFABP) gene, which is a marker for differentiated enterocytes. This is the first direct demonstration of a target for GATA factors in the vertebrate intestinal epithelium. Transactivation by xGATA-4, -5, and -6 is mediated at least in part by a defined proximal IFABP promoter element. The expression patterns for cGATA-4, -5, and -6 are markedly distinct along the proximal-distal villus axis. Transcript levels for cGATA-4 increase along the axis toward the villus tip; likewise, cGATA-5 transcripts are largely restricted to the distal tip containing differentiated cells. In contrast, the pattern of cGATA-6 transcripts is complementary to cGATA-5, with highest levels detected in the region of proliferating progenitor cells. Undifferentiated and proliferating human HT-29 cells express hGATA-6 but not hGATA-4 or hGATA-5. Upon stimulation to differentiate, the transcript levels for hGATA-5 increase, and this occurs prior to increased transcription of the terminal differentiation marker intestinal alkaline phosphatase. At the same time, hGATA-6 steady-state transcript levels decline appreciably. All of the data are consistent with evolutionarily conserved but distinct roles for these factors in regulating the differentiation program of intestinal epithelium. Based on this data, we suggest that GATA-6 might function primarily within the proliferating progenitor population, while GATA-4 and GATA-5 function during differentiation to activate terminal-differentiation genes including IFABP.

Identification and characterization of a novel collagenase in Xenopus laevis: possible roles during frog development.

Matrix metalloproteinases (MMPs) participate in extracellular matrix remodeling and degradation and have been implicated in playing important roles during organ development and pathological processes. Although it has been hypothesized for > 30 years that collagenase activities are responsible for collagen degradation during tadpole tail resorption, none of the previously cloned amphibian MMPs have been biochemically demonstrated to be collagenases. Here, we report a novel matrix metalloproteinase gene from metamorphosing Xenopus laevis tadpoles. In vitro biochemical studies demonstrate that this Xenopus enzyme is an interstitial collagenase and has an essentially identical enzymatic activity toward a collagen substrate as the human interstitial collagenase. Sequence comparison of this enzyme to other known MMPs suggests that the Xenopus collagenase is not a homologue of any known collagenases but instead represents a novel collagenase, Xenopus collagenase-4 (xCol4, MMP-18). Interestingly, during development, xCol4 is highly expressed only transiently in whole animals, at approximately the time when tadpole feeding begins, suggesting a role during the maturation of the digestive tract. More importantly, during metamorphosis, xCol4 is regulated in a tissue-dependent manner. High levels of its mRNA are present as the tadpole tail resorbs. Similarly, its expression is elevated during hindlimb morphogenesis and intestinal remodeling. In addition, when premetamorphic tadpoles are treated with thyroid hormone, the causative agent of metamorphosis, xCol4 expression is induced in the tail. These results suggest that xCol4 may facilitate larval tissue degeneration and adult organogenesis during amphibian metamorphosis.

Literate Language Intervention With High-Need Prekindergarten Children: A Randomized Trial.

Purpose: The present article reports on the implementation and results of a randomized intervention trial targeting the literate language skills of prekindergarten children without identified language disorders but with low oral language skills. Method: Children (N = 82; 45 boys and 37 girls) were screened-in and randomized to a business-as-usual control or to the pull-out treatment groups in which they received 4 instructional units addressing different sentence-level syntactic and semantic features: prepositions, conjunctions, adverbs, and negations. The intervention was delivered by paraprofessionals in small groups in the form of 20-min lessons 4 times a week for 12 weeks. Results: Overall, children receiving the supplemental instruction showed educationally meaningful gains in their oral language skills, relative to children in the control group. Significant group differences were found on researcher-designed oral language measures, with moderate to large effect sizes ranging from .44 to .88 on these measures. Conclusions: The intervention holds the potential to positively affect understanding and production of syntax and semantic features, such as prepositions and conjunctions, in young children with weak oral language skills.

Characterization of the Xenopus homolog of an immediate early gene associated with cell activation: sequence analysis and regulation of its expression by thyroid hormone during amphibian metamorphosis.

The complex transformation of a tadpole to a frog during amphibian development is under the control of thyroid hormone (T3). T3 is known to regulate gene transcription through its nuclear receptors. We have previously isolated many genes which are up-regulated by T3 in the intestine of Xenopus laevis tadpoles. We have now cloned a full-length cDNA for one such gene (IU12). Sequence analysis shows that the IU12 cDNA encodes a plasma membrane protein with 12 transmembrane domains and homologous to a mammalian gene associated with cell activation and organ development. Similarly, we have found that IU12 is activated during intestinal remodeling when both cell death and proliferation take place. Furthermore, IU12 is an early T3-response gene and its expression in the intestine during T3-induced metamorphosis mimics that during normal development. These results argue for a role of IU12 in the signal transduction pathways leading to intestinal metamorphosis.

Unique organization and involvement of GAGA factors in transcriptional regulation of the Xenopus stromelysin-3 gene.

Expression of the matrix metalloproteinase (MMP) gene stromelysin-3 ( ST3 ) has been shown to be tightly associated with cell migration and apoptosis inmammals and amphibians. This contrasts with most other MMP genes. We demonstrate here that the Xenopus ST3 gene also has a structure distinct from other MMP genes, with its C-terminal half (the hemopexin domain) encoded by 4 instead of 6 exons, as in other MMP genes. Our primer extension analysis reveals the existence of two transcription start sites and at least one is needed for transcription of the promoter in transient transfection assays. Furthermore, our deletion analysis has demonstrated a requirement for at least one GAGA factor binding site for promoter function. In vitro DNA binding and mutational studies have provided strong evidence for the participation of GAGA or GAGA-like factors in transcriptional regulation of the frog ST3 gene. This contrasts with regulation of the human ST3 promoter. These results suggest that the ST3 gene evolved prior to most other metalloproteinase genes and uses distinct regulation pathways to achieve similar expression profiles and serve similar functions in mammals and amphibians.