Patterns of Gene Expression in Microarrays and Expressed Sequence Tags from Normal and Cataractous Lenses

In this contribution, we have examined the patterns of gene expression in normal and cataractous lenses as presented in five different papers using microarrays and expressed sequence tags. The purpose was to evaluate unique and common patterns of gene expression during development, aging and cataracts

Conservation of the Three-Dimensional Structure in Non-Homologous Or Unrelated Proteins

In this review, we examine examples of conservation of protein structural motifs in unrelated or non-homologous proteins. For this, we have selected three DNA-binding motifs: the histone fold, the helix-turn-helix motif, and the zinc finger, as well as the globin-like fold. We show that indeed similar structures exist in unrelated proteins, strengthening the concept that three-dimensional conservation might be more important than the primary amino acid sequence

A Microarray Analysis of Gene Expression Patterns During Early Phases of Newt Lens Regeneration

Purpose: Notophthalmus viridescens, the red-spotted newt, possesses tremendous regenerative capabilities. Among the tissues and organs newts can regenerate, the lens is regenerated via transdifferentiation of the pigment epithelial cells of the dorsal iris, following complete removal (lentectomy). Under normal conditions, the same cells from the ventral iris are not capable of regenerating. This study aims to further understand the initial signals of lens regeneration

Gene expression during newt lens regeneration and cephalopod eye evolution

Newts are known for their ability to regenerate lost body parts. In contrast to many other organ systems, lens regeneration has many advantages. The eye lens can be removed as a whole and regeneration can occur through transdifferentiation of dorsal iris cells while ventral iris can be used as natural non-regenerating control. We have used microarrays, RNA-sequencing and mass spectrometry in dorsal and ventral iris samples during early phases of lens regeneration. The selected time points cover the undamaged control at 0 days post-lentecomy (dpl), the reentry of the cell cycle at 4 dpl and the beginning of transdifferentiation at 8 dpl. The newly assembled newt transcriptome was used to obtain annotation and gene expression measurements on newt genes in our samples. Functional analysis revealed genes related to redox balance, DNA repair, regulation of gene expression, cytoskeleton, immune response, metabolic processes, and cell cycle to be enriched in dorsal iris during regeneration time points. These events were associated with the transdifferentiation initiated in the dorsal iris. In addition, comparative transcriptomic and proteomic analyses using high-throughput gene expression data from other amphibian regeneration systems implicated response to stress, proliferation and migration, and cellular reprogramming to be a common program required for regeneration. Gene expression data from newt lens regeneration were extensively validated with quantitative real time polymerase chain reaction. Furthermore, microarrays in young and old axolotls, another amphibian model that was found capable of lens regeneration from the iris for a short window of two week after hatching, were used. Functional annotation indicated that young regeneration-competent axolotls expressed genes related to regulation of gene expression, electron transport chain, cell cycle, DNA repair and metabolic process -- gene groups belonging to the common regeneration program. In addition, we implicated immune response and cell differentiation in repression of lens regeneration in old axolotl iris. Cephalopods are protostome animals that exhibit an impressive vertebrate-like camera-type eye that facilitates high quality vision. Nautilus, however, has a pinhole eye that lacks cornea and lens. We used RNA-sequencing in developing Nautilus and pigmy squid embryos in order to gain more insights into cephalopod eye evolution. Pathway analysis of genes expressed only in Nautilus or pigmy squid developing eyes revealed that SIX3/6 gene is not expressed in the Nautilus. In addition, expression of all the genes regulated by this transcriptional factor was absent. Since, SIX3/6 is necessary for lens development in vertebrates and the gene network between vertebrates and invertebrates is highly conserved we argued that the absence of SIX3/6 in Nautilus leads to the pinhole eye. Functional and molecular evolution analyses of the Nautilus and pigmy squid transcriptomes revealed gene selections, and a gene duplication which might be associated with cephalopod eye evolution as well as in developing a vertebrate-like camera-type eye with invertebrate rhabdomere photoreceptors. The use of high-throughput methods in studying gene expression during newt lens regeneration and cephalopod eye evolution provided us with valuable insights into the underlying mechanisms in these systems

Gene expression during newt lens regeneration and cephalopod eye evolution

Newts are known for their ability to regenerate lost body parts. In contrast to many other organ systems, lens regeneration has many advantages. The eye lens can be removed as a whole and regeneration can occur through transdifferentiation of dorsal iris cells while ventral iris can be used as natural non-regenerating control. We have used microarrays, RNA-sequencing and mass spectrometry in dorsal and ventral iris samples during early phases of lens regeneration. The selected time points cover the undamaged control at 0 days post-lentecomy (dpl), the reentry of the cell cycle at 4 dpl and the beginning of transdifferentiation at 8 dpl. The newly assembled newt transcriptome was used to obtain annotation and gene expression measurements on newt genes in our samples. Functional analysis revealed genes related to redox balance, DNA repair, regulation of gene expression, cytoskeleton, immune response, metabolic processes, and cell cycle to be enriched in dorsal iris during regeneration time points. These events were associated with the transdifferentiation initiated in the dorsal iris. In addition, comparative transcriptomic and proteomic analyses using high-throughput gene expression data from other amphibian regeneration systems implicated response to stress, proliferation and migration, and cellular reprogramming to be a common program required for regeneration. Gene expression data from newt lens regeneration were extensively validated with quantitative real time polymerase chain reaction. Furthermore, microarrays in young and old axolotls, another amphibian model that was found capable of lens regeneration from the iris for a short window of two week after hatching, were used. Functional annotation indicated that young regeneration-competent axolotls expressed genes related to regulation of gene expression, electron transport chain, cell cycle, DNA repair and metabolic process -- gene groups belonging to the common regeneration program. In addition, we implicated immune response and cell differentiation in repression of lens regeneration in old axolotl iris. Cephalopods are protostome animals that exhibit an impressive vertebrate-like camera-type eye that facilitates high quality vision. Nautilus, however, has a pinhole eye that lacks cornea and lens. We used RNA-sequencing in developing Nautilus and pigmy squid embryos in order to gain more insights into cephalopod eye evolution. Pathway analysis of genes expressed only in Nautilus or pigmy squid developing eyes revealed that SIX3/6 gene is not expressed in the Nautilus. In addition, expression of all the genes regulated by this transcriptional factor was absent. Since, SIX3/6 is necessary for lens development in vertebrates and the gene network between vertebrates and invertebrates is highly conserved we argued that the absence of SIX3/6 in Nautilus leads to the pinhole eye. Functional and molecular evolution analyses of the Nautilus and pigmy squid transcriptomes revealed gene selections, and a gene duplication which might be associated with cephalopod eye evolution as well as in developing a vertebrate-like camera-type eye with invertebrate rhabdomere photoreceptors. The use of high-throughput methods in studying gene expression during newt lens regeneration and cephalopod eye evolution provided us with valuable insights into the underlying mechanisms in these systems

NF1/Nf1 sequence comparisons.

<p>A. Nucleotide sequence alignment using ClustalW2 of squid contigs that were found from this analysis indicate potential gene duplication of NF1/Nf1 in cephalopods. Comp304995_c0_seq1 has best hit with Drosophila Nf1 and comp475299_c0_seq1 with Human NF1. Asterisk(*) indicate sequence identity. B. Phylogenetic tree using Human NF1 and NF2, Drosophila Nf1 and Merlin as well as <i>Nautilus</i> and squid NF1/Nf1 contigs. Note that Nautilus and squid NF1 contigs, and Nautilus and squid Nf1 contigs cluster together. Bar indicates phylogenetic distance.</p

An example of Human/Drosophila conserved protein.

<p>PPP2CA/mts pair was found in <i>Nautilus</i> and squid as a potential candidate for lens evolution. These pairs were discarded from our analysis due to the high conservation of the Human and Drosophila proteins. Asterisk(*) indicate sequence identity, colon(:) indicates strongly similar properties and period(.) indicates weakly similar properties (ClustalW2).</p

Alignments of the three gene pairs found to be selected for lens evolution in squids.

<p>Drosophila and Human homologue protein sequence were aligned using ClustalW2 to show conserved regions. Highlighted areas indicate the squid or <i>Nautilus</i> hit sequences with their counterpart homologues in Human or Drosophila. Proteins are labeled with their UniProt entry. A. CAP1/capt gene pair alignment. Squid has a best hit with the Human homologue (CAP1; CAP1_HUMAN) while <i>Nautilus</i> has a best hit with the Drosophila homologue (capt; Q9VPX7_DROME) in the same region. B. RAPGEF2/Gef26 gene pair alignment. Squid has a best hit with the Drosophila homologue (Gef26; Q9VMF3_DROME) while <i>Nautilus</i> has a best hit with the Human homologue (RAPGEF2; RPGF2_HUMAN) in the same region. C. CD2BP2/CG5198 gene pair alignment. Squid has a best hit with the Drosophila homologue (CG5198; LIN1_DROME) while <i>Nautilus</i> has a best hit with the Human homologue (CD2BP2; CD2B2_HUMAN) in the same region. Asterisk(*) indicate sequence identity, colon(:) indicates strongly similar properties and period(.) indicates weakly similar properties (ClustalW2).</p