Identification of Genes Related to Beak Deformity of Chickens Using Digital Gene Expression Profiling

<div><p>Frequencies of up to 3% of beak deformity (normally a crossed beak) occur in some indigenous chickens in China, such as and Beijing-You. Chickens with deformed beaks have reduced feed intake, growth rate, and abnormal behaviors. Beak deformity represents an economic as well as an animal welfare problem in the poultry industry. Because the genetic basis of beak deformity remains incompletely understood, the present study sought to identify important genes and metabolic pathways involved in this phenotype. Digital gene expression analysis was performed on deformed and normal beaks collected from Beijing-You chickens to detect global gene expression differences. A total of >11 million cDNA tags were sequenced, and 5,864,499 and 5,648,877 clean tags were obtained in the libraries of deformed and normal beaks, respectively. In total, 1,156 differentially expressed genes (DEG) were identified in the deformed beak with 409 being up-regulated and 747 down-regulated in the deformed beaks. qRT-PCR using eight genes was performed to verify the results of DGE profiling. Gene ontology (GO) analysis highlighted that genes of the keratin family on GGA25 were abundant among the DEGs. Pathway analysis showed that many DEGs were linked to the biosynthesis of unsaturated fatty acids and glycerolipid metabolism. Combining the analyses, 11 genes (<i>MUC</i>, <i>LOC426217</i>, <i>BMP4</i>, <i>ACAA1</i>, <i>LPL</i>, <i>ALDH7A1</i>, <i>GLA</i>, <i>RETSAT</i>, <i>SDR16C5</i>, <i>WWOX</i>, and <i>MOGAT1</i>) were highlighted as potential candidate genes for beak deformity in chickens. Some of these genes have been identified previously, while others have unknown function with respect to thus phenotype. To the best of our knowledge, this is the first genome-wide study to investigate the transcriptome differences in the deformed and normal beaks of chickens. The DEGs identified here are worthy of further functional characterization.</p></div

Sequencing saturation analysis for RNA from the deformed (A) and normal beak (B).

<p>Sequencing saturation analysis for RNA from the deformed (A) and normal beak (B).</p

The deformed (left) and normal (right) beaks of Beijing-You chickens.

<p>The chicken with a deformed beak had problem with feeding, drinking, and preening and therefore showed lower body weight and poor mental condition.</p

The two most significantly enriched pathways and the involved differentially expressed genes (DEG).

a<p><i>SCD5</i>  =  stearoyl-CoA desaturase 5; <i>GLG1</i>  =  golgi glycoprotein 1; <i>ELOVL6</i>  =  ELOVL family member 6; <i>PTPLB</i>  =  protein tyrosine phosphatase-like B; <i>ACAA1</i>  =  acetyl-CoA acyltransferase 1; <i>PECR</i>  =  privacy and electronic communications regulations; <i>MFSD4</i>  =  major facilitator superfamily domain containing 4; <i>DLEC1</i>  =  deleted in lung and esophageal cancer 1; <i>PTPLAD1</i>  =  protein tyrosine phosphatase-like A domain containing 1; <i>HSDL1</i>  =  hydroxysteroid dehydrogenase like gene; <i>LOC423119</i>  =  Gallus gallus fatty acid desaturase 1-like; <i>LPL</i>  =  lipoprotein lipase; <i>PNPLA2</i>  =  patatin-like phospholipase domain containing 2; <i>PPAP2A</i>  =  phosphatidic acid phosphatase 2A; <i>PPAP2B</i>  =  phosphatidic acid phosphatase 2B; <i>AGPAT6</i>  =  1-acylglycerol-3-phosphate O-acyltransferase 6; <i>SHROOM3</i>  =  shroom family member 3; <i>ALDH7A1</i>  =  aldehyde dehydrogenase 7 family, member A1; <i>AKR1B10</i>  =  aldo-keto reductase family 1, member B10; <i>DGKD</i>  =  diacylglycerol kinase; <i>GK5</i>  =  glycerol kinase 5; <i>GLA</i>  =  galactosidase, alpha; <i>MOGAT1</i>  =  monoacylglycerol O-acyltransferase 1; <i>AR</i>  =  androgen receptor; <i>LIPG</i>  =  endothelial lipase.</p><p>The two most significantly enriched pathways and the involved differentially expressed genes (DEG).</p

Gene-specific primers used in qRT-PCR.

a<p><i>NPM3</i>  =  nucleophosmin/nucleoplasmin 3; <i>LPL</i>  =  lipoprotein lipase; <i>BMP4</i>  =  bone morphogenetic protein 4; <i>SGOL1</i>  =  shugoshin-like 1; <i>KRT19</i>  =  Keratin 19; <i>sKer</i>  =  similar to Scale keratin; <i>MMP7</i>  =  matrix metalloproteinase 7; <i>MUC</i>  =  mucin protein.</p><p>Gene-specific primers used in qRT-PCR.</p

Top 10 enrichment cellular components.

<p>The numbers of genes involved in the components are indicated in parentheses.</p

Top 10 enrichment pathways.

<p>The numbers of genes involved in the pathways are indicated in parentheses.</p

Additional file 2: of Study on LOC426217 as a candidate gene for beak deformity in chicken

The ARRIVE Guidelines Checklist Animal Research: Reporting In Vivo Experiments. (DOCX 661 kb

Some extremely differentially expressed genes (|log2-Ratio (deformed beak/normal beak)| ≧ 9).

a<p><i>LOC426217</i>  =  claw keratin-like; <i>NPM3</i>  =  nucleophosmin/nucleoplasmin 3; <i>LPL</i>  =  lipoprotein lipase; <i>RBP7</i>  =  retinol binding protein 7 cellular; <i>NUBP2</i>  =  nucleotide binding protein 2 (MinD homolog <i>E. coli</i>); <i>ARL6IP1</i>  =  ADP-ribosylation factor-like 6 interacting protein 1; <i>ABF1  =  activated B-cell factor 1</i>; <i>RNG213</i>  =  ring finger protein 213; <i>THOC3</i>  =  THO complex 3; <i>DGCR14</i>  =  DiGeorge syndrome critical region gene 14; <i>C3orf38</i>  =  chromosome 3 open reading frame 38; <i>KRT19</i>  =  Keratin 19; <i>SGOL1</i>  =  shugoshin-like 1; <i>sKer</i>  =  similar to Scale keratin; <i>MMP7</i>  =  matrix metalloproteinase 7; <i>MUC</i> =  mucin protein.</p><p>Some extremely differentially expressed genes (|log2-Ratio (deformed beak/normal beak)| ≧ 9).</p

qRT-PCR of 8 transcripts for validating the DGE results.

<p>The horizontal axis identifies the 8 transcripts examined by qRT-PCR; the vertical axis shows the relative gene expression level in deformed (individuals 1 and 2) versus normal (individuals 3, 4, 5, and 6) beak tissues. The 4 pairs of individuals were all full sibs. The first bar shows the value obtained from DEG using 1 and 6. <i>NPM3</i>  =  nucleophosmin/nucleoplasmin 3; <i>LPL</i>  =  lipoprotein lipase; <i>BMP4</i>  =  bone morphogenetic protein 4; <i>SGOL1</i>  =  shugoshin-like 1; <i>KRT19</i>  =  Keratin 19; <i>sKer</i>  =  similar to Scale keratin; <i>MMP7</i>  =  matrix metalloproteinase 7; <i>MUC</i>  =  mucin protein.</p