Distinct Developmental Expression Patterns Research Articles

Molecular cloning and distinct developmental expression pattern of spliced forms of a novel zinc finger gene wiz in the mouse cerebellum

In the course of a study conducted to identify the mouse homologue of Drosophila eyes absent ( eya), we isolated a novel mouse cDNA fragment which show little homology to eya but encodes a protein with Krüppel (C 2H 2)-type zinc finger motifs. By further screening using this cDNA fragment as a probe, we obtained the short and long forms of full-length cDNAs, which were apparently alternatively spliced products from one gene. Since both mRNAs encode proteins with widely- interspaced zinc finger motifs, we termed this gene wiz and refer to the short and long wiz transcripts as wizS and wizL, respectively. In situ hybridization studies using the probe against the region common to wizS and wizL showed that these mRNAs were expressed abundantly in the granule cell layers of the mouse cerebellum, the olfactory bulb, and the dentate gyrus, whereas the same technique using the probe against only wizL could not detect positive signals in the developing cerebellum, indicating that there is no expression of wizL mRNA there. Northern blot and in situ hybridization analyses demonstrated that the extracerebellar regions expressed both wizS and wizL mRNAs from the midgestational period to adulthood. The finding that two types of wiz transcripts ( wizS and wizL) are expressed with different developmental patterns might indicate separate transcription functions in the cerebellar granule cells and the extracerebellar regions.

Proenkephalin gene expression in testicular interstitial cells is down-regulated coincident with the appearance of pachytene spermatocytes.

Two distinct forms of proenkephalin messenger RNA (mRNA) are present in the murine testis, a family of 1.7 kilobases (kb), germ cell-specific transcripts and a 1.45-kb form that is also found in somatic tissues. In situ hybridization and molecular analysis of purified spermatogenic cell types were used to characterize the cellular localization of these different transcripts during development of the mouse testis. Both forms of proenkephalin mRNA were observed in isolated germ cells by RNA gel-blot analysis, but in distinct developmental patterns; the 1.7-kb transcripts were present in cells undergoing meiosis and spermiogenesis, whereas the 1.45-kb mRNA was detected primarily in type B spermatogonia. In contrast, in situ hybridization analysis did not detect significant amounts of the 1.45-kb transcript in any spermatogenic cell type. Using transcript-specific probes, distinct patterns of developmental expression were evident for the two mRNAs. The 1.45-kb transcript was the only form detected in the prepubertal testis, where it was localized mainly in interstitial cells. In contrast, the 1.7-kb transcripts were the major mRNAs observed in the adult testis and were localized to spermatogenic cells. A transition from the prepubertal to the adult pattern occurred on or about postnatal day 21, when proenkephalin-expressing pachytene spermatocytes begin to populate the seminiferous tubules. In situ hybridization analysis further demonstrated that proenkephalin gene expression in mutant (at/at) mice, which lack germ cells, was identical to that observed in the early prepubertal testis. These results suggest that the 1.45-kb proenkephalin mRNA is developmentally down-regulated in mouse interstitial cells and that this process requires ongoing spermatogenesis.

AE3 anion exchanger isoforms in the vertebrate retina: developmental regulation and differential expression in neurons and glia

Plasma membrane anion exchangers constitute a multigene family that contributes to the regulation of intracellular pH and chloride concentration in many cell types. We have characterized two polypeptide isoforms of the AE3 gene that are expressed in the rat retina. Using antipeptide antibodies specific for defined NH2-terminal and COOH-terminal epitopes, we have identified a 165 kDa polypeptide whose expression is restricted to the primary glial cell type of the retina, the Müller cell, and a 125 kDa polypeptide that is expressed in horizontal neurons. Expression of the Müller cell isoform exhibits a polarized distribution and is highest in basal endfoot processes. These AE3 isoforms exhibit a distinct developmental expression pattern in postnatal rat retina. The neuronal isoform is undetectable in neonatal retina until postnatal day 10-15, correlating strongly with the onset of retinal function.

Coordinate regulation of tubulin and microtubule associated protein genes during development of hamster brain

The present study documents the patterns of mRNA expression for 5 different tubulin genes and 4 of the structural microtubule-associated protein (MAP) genes during normal development of hamster forebrain. Northern blotting in conjunction with densitometric analysis was used to study changes in the levels of the mRNAs for α I-tubulin, classes β I-, β II-, β III- and β IV-tubulin, as well as the mRNAs for tau, MAP1A, MAP1B and MAP2, using total RNA isolated from hamster forebrain at various embryonic (E) and postnatal (P) stages. Densitometric analyses of the autoradiograms from the Northern blots revealed that each of the tubulin genes exhibited distinct developmental patterns of expression, several of which appeared to be temporally correlated with the expression of specific MAP mRNAs. The β I-, β II- and β III-tubulin mRNAs increased rapidly between late embryonic stages to birth, reached peak levels early in the first postnatal week, and declined thereafter. α I-Tubulin mRNA was easily detected during embryonic stages, rose to peak levels at P7–P9 and then gradually declined. A similar pattern was seen for tau mRNA. After the first postnatal week, the size of the tau mRNA also shifted to a slightly larger size, presumably due to differential splicing. β IV-Tubulin mRNA levels did not become significant until very late in postnatal development (3–4 weeks). MAP2 mRNA expression was unusual in that peak levels were reached at two different stages of development - an initial peak occurred in the first postnatal week, followed by a decline, and then a second rise occurred during the third and fourth postnatal weeks. The expression of the β IV-tubulin mRNA coincided temporally with the second peak in MAP2 mRNA expression. MAP1B mRNA abruptly reached high levels at birth, remained abundant during the first two postnatal weeks, and then decreased. In contrast, MAP1A mRNA levels were low in the initial postnatal interval and increased only at very late developmental stages. The findings of a temporal correspondence in expression of high levels of tau and MAP1B with β I-, β II-, β III- and αI-tubulin mRNAs suggest that this profile of gene expression is one that endows greater plasticity to the neuronal cytoskeleton. Conversely, the transition to increased expression of β IV-tubulin, MAP1A, and a larger tau mRNA species defines a portion of the molecular pattern that underpins the increased stability of neuronal form during maturation.

Closely related DNA sequences specify distinct patterns of developmental expression in Drosophila melanogaster.

Three short synthetic DNA sequences, which are closely related to one another, confer three distinct patterns of developmental expression on the heat shock hsp70 gene in transgenic Drosophila melanogaster lines. These results show that small variations or even single base pair changes in a repeated element of a regulatory sequence can create promoters that display new specificities of tissue and developmental regulation. Interestingly, the three patterns of developmental expression conferred by the synthetic DNAs resemble in part those of the known developmental genes: glucose dehydrogenase (Gld), Dopa decarboxylase (Ddc), and salivary gland secretory proteins (Sgs), respectively. In each case, the defined regulatory region of the known developmental gene contains multiple sequences that are similar or identical to the synthetic sequence that confers a similar pattern of developmental expression on the hsp70 gene. Thus, these results are congruent with the view that short sequence elements in multiple copies can confer either simple or relatively complex patterns of developmental expression on a receptive promoter like that of hsp70. Furthermore, the fact that the three variants tested produced three distinct patterns of expression in transgenic animals suggests that the number of different elements is large.

Closely Related DNA Sequences Specify Distinct Patterns of Developmental Expression in Drosophila melanogaster

Three short synthetic DNA sequences, which are closely related to one another, confer three distinct patterns of developmental expression on the heat shock hsp70 gene in transgenic Drosophila melanogaster lines. These results show that small variations or even single base pair changes in a repeated element of a regulatory sequence can create promoters that display new specificities of tissue and developmental regulation. Interestingly, the three patterns of developmental expression conferred by the synthetic DNAs resemble in part those of the known developmental genes: glucose dehydrogenase (Gld), Dopa decarboxylase (Ddc), and salivary gland secretory proteins (Sgs), respectively. In each case, the defined regulatory region of the known developmental gene contains multiple sequences that are similar or identical to the synthetic sequence that confers a similar pattern of developmental expression on the hsp70 gene. Thus, these results are congruent with the view that short sequence elements in multiple copies can confer either simple or relatively complex patterns of developmental expression on a receptive promoter like that of hsp70. Furthermore, the fact that the three variants tested produced three distinct patterns of expression in transgenic animals suggests that the number of different elements is large.

Developmental Regulation of Calmodulin, Actin, and Tubulin Rnas during Rat Testis Differentiation1

Postnatal testis differentiation involves transition through neonatal, pre-meiotic, meiotic, haploid, and mature stages. We have examined the qualitative and quantitative changes in rat testis RNAs that specifically hybridize to cDNAs encoding the cytoskeletal proteins, calmodulin, beta-actin, alpha- and beta-tubulin at ages corresponding to each of these developmental periods. We compared the species and relative levels of specific RNAs from testes of animals engaged in normal spermatogenesis with RNA from germ cell-depleted, Sertoli cell-enriched (SCE) testis. Distinct developmental patterns of expression of the specific RNAs were found with each of the cDNAs in the two animal models. A 2.2 kb (kilobase) actin RNA and a 2.7 kb beta-tubulin RNA are maximal at 5-10 days of age, suggesting these RNAs are required by somatic and germ cells in the postnatal phase prior to puberty. Between 19 and 29 days, when pachytene spermatocytes appear in significant numbers, there is a slight increase in the 2.2-kb actin RNA, but a 4- to 10-fold increase in RNAs hybridizing to cDNAs for calmodulin, alpha- and beta-tubulin. These changes are much less pronounced in the SCE testis than in the normal testis, indicating increases in these RNAs are related to germinal cell maturation. The germ cell-related increase in 1.8-kb beta-tubulin RNA appears to reflect a developmental "switch" in the gene from which the RNA is derived. This hypothesis is based on the observation that the ratio of hybridization of a chicken brain beta-tubulin cDNA versus a rat spleen beta-tubulin cDNA to the 1.8-kb RNA band increases more than 40-fold between 5 and 29 days of age in normal testis, but is constant in SCE testis. These data suggest that a specific beta-tubulin gene is activated in maturing germ cells. Analogously, a 2.1-kb alpha-tubulin RNA is found only in maturing normal testis and increases as spermatids are produced. A 2.0-kb beta-tubulin RNA, not found in normal testes, is maximal in maturing SCE testes, suggesting this RNA is of somatic cell origin. All of the RNA species studied, except the 2.0-kb beta-tubulin RNA, decrease between 5 and 19 days in SCE testes, as Sertoli cell mitotic activity wanes, indicating that their levels may be regulated by the developmental signals that influence mitosis.