XB-ART-4532Mech Dev October 1, 2003; 120 (10): 1177-92.
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Developmental role of HMGN proteins in Xenopus laevis.
HMGN proteins are architectural chromatin proteins that reduce the compaction of the chromatin fiber, facilitate access to nucleosomes and modulate replication and transcription processes. Here we demonstrate that in Xenopus laevis, the expression and cellular location of the HMGN proteins are developmentally regulated and that their misexpression leads to gross developmental defects in post-blastula embryos. HMGN transcripts and proteins are present throughout oogenesis; however, the proteins stored in the cytoplasm are not associated with lampbrush chromosomes, and are rapidly degraded when oocytes mature into eggs. During embryogenesis, HMGN expression is first detected in blastula stages and progresses to a tissue-specific expression reaching relative high levels in the mesodermal and neuroectodermal regions of tadpoles. Only after midblastula transition (MBT), alterations in the HMGN levels by either microinjection of recombinant proteins or by morpholino-antisense oligo treatments produced embryos with imperfectly closed blastopore, distorted body axis and showed abnormal head structures. Analyses of animal cap explants indicated that HMGN proteins are involved in the regulation of mesoderm specific genes. In addition, HMGN misexpression caused altered expression of specific genes at MBT rather than global changes of transcription rates. Our results demonstrate that proper embryonic development of Xenopus laevis requires precisely regulated levels of HMGN proteins and suggest that these nucleosomal binding proteins modulate the expression of specific genes.
PubMed ID: 14568106
Article link: Mech Dev
Species referenced: Xenopus laevis
Genes referenced: hmgn1 hmgn2 myod1 nrp1
Morpholinos: hmgn2 MO1
Article Images: [+] show captions
|Fig. 4. Expression pattern of HMGN genes during Xenopus embryogenesis. (A) Northern blot analysis was performed as described in Fig. 2. In each lane total RNA of five embryos each of following stages was analyzed: 2-cell, 4-cell, 16-cell embryo, early blastula (stage 6, 5), blastula (stage 8), gastrula (stage 11), neurula (stage 19), stage 25/26 embryos. As a control for the amount of RNA loaded, ethidium bromide staining of the 18S ribosomal RNA is shown. XHMGN1 and XHMGN2 transcripts are first detectable at the blastula stage. (B) Whole-mount in situ hybridization experiments showing the localization of XHMGN1 (a–d) and XHMGN2 (e, f) transcripts in neurula (a, c, e) and tailbud (b, d, f) embryos. Bright field illumination of BB/BA cleared embryos is shown in (a, b) and top illumination of uncleared embryos in (c–f), respectively. HMGN1/N2 genes are expressed in all tissues but are particularly abundant in mesodermal and neuroectodermal regions of the embryos. Abbreviations: ncr, neural crest; s, somite; cg, cement gland; ant, anterior; post, posterior. Bar represents 0.5 mm in (a) and 1.3 mm in (b). (C) NF25/26 embryos were dissected as indicated in the cartoon into mesoderm/neural tube (3, red), brain/head (2, yellow), endoderm (4, green) and ectoderm/skin (1). RNA was isolated from dissected tissues and probed for HMGN1 or HMGN2 expression by RT-PCR. The expression of the pan-neural marker NRP-1 and the mesodermal marker XmyoD were used to show the enrichment of respective tissues in the embryonic fractions. As a control, the expression of the constitutively expressed translation elongation factor EF1-α is shown. Note the elevated expression levels of HMGN1 and HMGN2 in neural and mesodermal tissues.|