Molecular basis for temporal patterns of sea urchin embryonic gene expression

Abstract

In sea urchins, there are at least two classes of histone genes that are differentially expressed during early embryogenesis. The early genes are maximally transcribed at the early blastula state and inactivated during later development. In contrast, the late genes are activated in late blastula and gastrula and in adult tissues. The objective of this research is to understand the molecular mechanisms for the differential expression of these genes during early embryogenesis.;To approach this question, early H1-{dollar}\alpha{dollar}, early H3 and late H1-{dollar}\beta{dollar} genes were studied in detail by using an in vivo transient expression system involving microinjection of cloned DNA constructs into fertilized eggs. Following injection, each of these genes are expressed at the appropriate time during embryogenesis, indicating that the injected DNA molecules contain all the information needed for correct regulation. Both positive and negative cis-acting DNA elements were identified in the early H3 gene promoter. In the case of late H1-{dollar}\beta{dollar} gene, multiple regulatory elements were identified in the upstream sequences by using 5{dollar}\sp\prime{dollar} deletion, internal deletion, and point mutation constructs. A sequence of 30 bp (USE4) located between {dollar}-317{dollar} and {dollar}-288{dollar} has been identified as an embryonic stage-specific enhancer capable of activating heterologous promoters at the last blastula stage. The temporal pattern can be affected by mutations in the basal elements of the promoter as well. Furthermore, in vivo competition experiments demonstrate the existence of positive trans-acting factors which interact with the H1-{dollar}\beta{dollar} gene enhancer and basal promoter elements. Thus, embryonic stage-specific enhancer, basal promoter elements, and trans-acting factors, determine the timing and level of expression of the late H1-{dollar}\beta{dollar} gene during early embryogenesis. As expected, the sea urchin sperm H2B-2 gene, which is tissue-specifically expressed only in male testis, is inactive in the injected embryos.