Conservation of gene linkage in dispersed vertebrate NK homeobox clusters

Nk homeobox genes are important regulators of many different developmental processes including muscle, heart, central nervous system and sensory organ development. They are thought to have arisen as part of the ANTP megacluster, which also gave rise to Hox and ParaHox genes, and at least some NK genes remain tightly linked in all animals examined so far. The protostome–deuterostome ancestor probably contained a cluster of nine Nk genes: (Msx)–(Nk4/tinman)–(Nk3/bagpipe)–(Lbx/ladybird)–(Tlx/c15)–(Nk7)–(Nk6/hgtx)–(Nk1/slouch)–(Nk5/Hmx). Of these genes, only NKX2.6–NKX3.1, LBX1–TLX1 and LBX2–TLX2 remain tightly linked in humans. However, it is currently unclear whether this is unique to the human genome as we do not know which of these Nk genes are clustered in other vertebrates. This makes it difficult to assess whether the remaining linkages are due to selective pressures or because chance rearrangements have “missed” certain genes. In this paper, we identify all of the paralogs of these ancestrally clustered NK genes in several distinct vertebrates. We demonstrate that tight linkages of Lbx1–Tlx1, Lbx2–Tlx2 and Nkx3.1–Nkx2.6 have been widely maintained in both the ray-finned and lobe-finned fish lineages. Moreover, the recently duplicated Hmx2–Hmx3 genes are also tightly linked. Finally, we show that Lbx1–Tlx1 and Hmx2–Hmx3 are flanked by highly conserved noncoding elements, suggesting that shared regulatory regions may have resulted in evolutionary pressure to maintain these linkages. Consistent with this, these pairs of genes have overlapping expression domains. In contrast, Lbx2–Tlx2 and Nkx3.1–Nkx2.6, which do not seem to be coexpressed, are also not associated with conserved noncoding sequences, suggesting that an alternative mechanism may be responsible for the continued clustering of these genes.     

Isoform-specific O-glycosylation by murine UDP-GalNAc:polypeptide N- acetylgalactosaminyltransferase-T3, in vivo

Multiple isoforms of UDP-GalNAc:polypeptide N-acetylgalactosaminyl- transferase (ppGaNTase) have been cloned and expressed from a variety of organisms. In general, these isoforms display different patterns of tissue-specific expression, but exhibit overlapping substrate specificities, in vitro . A peptide substrate, derived from the sequence of the V3 loop of the HIV gp120 protein (HIV peptide), has previously been shown to be glycosylated in vitro exclusively by the ppGaNTase-T3 (Bennett et al. , 1996). To determine if this isoform- specificity is maintained in vivo , we have examined the glycosylation of this substrate when it is expressed as a reporter peptide (rHIV) in a cell background (COS7 cells) which lacks detectable levels of the ppGaNTase-T3. Glycosylation of rHIV was greatly increased by coexpression of a recombinant ppGaNTase-T3. Overexpression of ppGaNTase- T1 yielded only partial glycosylation of the reporter. We have also determined that the introduction of a proline residue at the +3 position flanking the potential glycosylation site eliminated ppGaNTase- T3 selectivity toward rHIV observed both in vivo and in vitro .      

Comparative genomics of Lbx loci reveals conservation of identical Lbx ohnologs in bony vertebrates

Background  

Lbx/ladybird genes originated as part of the metazoan cluster of Nk homeobox genes. In all animals investigated so far, both the protostome genes and the vertebrate Lbx1 genes were found to play crucial roles in neural and muscle development. Recently however, additional Lbx genes with divergent expression patterns were discovered in amniotes. Early in the evolution of vertebrates, two rounds of whole genome duplication are thought to have occurred, during which 4 Lbx genes were generated. Which of these genes were maintained in extant vertebrates, and how these genes and their functions evolved, is not known.