Mammary transcriptome reveals cell maintenance and protein turnover support milk synthesis in early-lactation cows.

A more complete understanding of the molecular mechanisms that support milk synthesis is needed to develop strategies to efficiently and sustainably meet the growing global demand for dairy products. With the postulate that coding gene transcript abundance reflects relative importance in supporting milk synthesis, we analyzed the global transcriptome of early lactation cows across magnitudes of normalized RNA-Seq read counts. Total RNA was isolated from milk samples collected from early-lactation cows (n = 6) following two treatment periods of postruminal lysine infusion of 0 or 63 g/day. Twelve libraries were prepared and sequenced on an Illumina NovaSeq6000 platform using paired end reads. Normalized read counts were averaged across both treatments, because EBseq analysis found no significant effect of lysine infusion. Approximately 10% of the total reads corresponded to 12,730 protein coding transcripts with a normalized read count mean ≥5. For functional annotation analysis, the protein coding transcripts were divided into nine categories by magnitude of reads. The 13 most abundant transcripts (≥50K reads) accounted for 67% of the 23M coding reads and included casein and whey proteins, regulators of fat synthesis and secretion, a ubiquitinating protein, and a tRNA transporter. Mammalian target of rapamycin, JAK/STAT, peroxisome proliferator-activated receptor alpha, and ubiquitin proteasome pathways were enriched with normalized reads ≥100 counts. Genes with ≤100 reads regulated tissue homeostasis and immune response. Enrichment in ontologies that reflect maintenance of translation, protein turnover, and amino acid recycling indicated that proteostatic mechanisms are central to supporting mammary function and primary milk component synthesis.

Global transcriptional differences in myokine and inflammatory genes in muscle of mature steer progeny are related to maternal lactation diet and muscle composition.

Steer progeny suckled by cows fed a dried distillers grains and solubles (DDGS) diet the first 3 mo of lactation were heavier during feedlot finishing and had significantly lower marbling and larger longissimus muscles than steers suckled by cows fed a control diet (CON). These differences were profound in that progeny were managed and fed identically from weaning until finishing, and findings suggest that the suckling period established the developmental program of muscle composition. Here transcriptomes of longissimus muscle were measured by next-generation sequencing to investigate whether there were any developmental clues to the differences in marbling scores and muscle content between steers suckled by DDGS ( n = 5) vs. control (CON; n = 5) diet-fed cows during lactation. There were 809 genes differentially expressed ( P-adj<0.1) between CON and DDGS muscle. Of these 636 were upregulated and 173 downregulated in DDGS relative to CON. Overall the DDGS vs. CON muscle transcriptomic signature was promyogenic and antiadipogenic. In particular, myokines/satellite cell maintenance factors were found among upregulated (LIF, CNTF, FGFB1, EPHB1) genes. The antiadipogenic signature was typified by the upregulation of anti-inflammatory cytokines and receptors (IL1RAP, IL1RL2, IL13RA2, IL1F10), and downregulation of expression of inflammation/inflammatory cytokines and receptor (TNF, IL6R, CXCL9), which suggests a selection of differentiation pathways away from adipogenic line. The upregulation of TGFB, SPP1, and INHBA supports selection of fibroblast lineage of cells. Thus, the lactation phase of production can effect meat quality by affecting transcriptional signatures that favor myogenesis and depress inflammation.

Effect of high-fat diet on secreted milk transcriptome in midlactation mice.

High-fat diet (HFD) during lactation alters milk composition and is associated with development of metabolic diseases in the offspring. We hypothesized that HFD affects milk microRNA (miRNA) and mRNA content, which potentially impact offspring development. Our objective was to determine the effect of maternal HFD on secreted milk transcriptome. To meet this objective, 4 wk old female ICR mice were divided into two treatments: control diet containing 10% kcal fat and HFD containing 60% kcal fat. After 4 wk on CD or HFD, mice were bred while continuously fed the same diets. On postnatal day 2 (P2), litters were normalized to 10 pups, and half the pups in each litter were cross-fostered between treatments. Milk was collected from dams on P10 and P12. Total RNA was isolated from milk fat fraction of P10 samples and used for mRNA-Seq and small RNA-Seq. P12 milk was used to determine macronutrient composition. After 4 wk of prepregnancy feeding HFD mice weighed significantly more than did the control mice. Lactose and fat concentration were significantly ( P < 0.05) higher in milk of HFD dams. Pup weight was significantly greater ( P < 0.05) in groups suckled by HFD vs. control dams. There were 25 miRNA and over 1,500 mRNA differentially expressed (DE) in milk of HFD vs. control dams. DE mRNA and target genes of DE miRNA enriched categories that were primarily related to multicellular organismal development. Maternal HFD impacts mRNA and miRNA content of milk, if bioactive nucleic acids are absorbed by neonate differences may affect development.

Channel catfish BAC-end sequences for marker development and assessment of syntenic conservation with other fish species.

In the present study, 25 195 BAC ends for channel catfish (Ictalurus punctatus) were sequenced, generating 20 366 clean BAC-end sequences (BES), with an average read length of 557 bp after trimming. A total of 11 414 601 bp were generated, representing approximately 1.2% of the catfish genome. Based on this survey, the catfish genome was found to be highly AT-rich, with 60.7% A+T and 39.3% G+C. Approximately 12% of the catfish genome consisted of dispersed repetitive elements, with the Tc1/mariner transposons making up the largest percentage by base pair (4.57%). Microsatellites were detected in 17.5% of BES. Catfish BACs were anchored to the zebrafish and Tetraodon genome sequences by BLASTN, generating 16% and 8.2% significant hits (E < e(-5)) respectively. A total of 1074 and 773 significant hits were unique to the zebrafish and Tetraodon genomes, respectively, of which 417 and 406, respectively, were identified as known genes in other species, providing a major genome resource for comparative genomic mapping.

Genes expressed during the development and ripening of watermelon fruit.

A normalized cDNA library was constructed using watermelon flesh mRNA from three distinct developmental time-points and was subtracted by hybridization with leaf cDNA. Random cDNA clones of the watermelon flesh subtraction library were sequenced from the 5' end in order to identify potentially informative genes associated with fruit setting, development, and ripening. One-thousand and forty-six 5'-end sequences (expressed sequence tags; ESTs) were assembled into 832 non-redundant sequences, designated as "EST-unigenes". Of these 832 "EST-unigenes", 254 ( approximately 30%) have no significant homology to sequences published so far for other plant species. Additionally, 168 "EST-unigenes" ( approximately 20%) correspond to genes with unknown function, whereas 410 "EST-unigenes" ( approximately 50%) correspond to genes with known function in other plant species. These "EST-unigenes" are mainly associated with metabolism, membrane transport, cytoskeleton synthesis and structure, cell wall formation and cell division, signal transduction, nucleic acid binding and transcription factors, defense and stress response, and secondary metabolism. This study provides the scientific community with novel genetic information for watermelon as well as an expanded pool of genes associated with fruit development in watermelon. These genes will be useful targets in future genetic and functional genomic studies of watermelon and its development.

Characterization and expression of beta-1,3-glucanase genes in peach.

Beta-1,3-glucanase is one of the pathogenesis-related (PR) proteins involved in plant defense responses. A peach beta-1,3-glucanase gene, designated PpGns1, has been isolated and characterized. The deduced amino acid sequence of the product of PpGns indicates that it is a basic isoform (pI 9.8), and contains a putative signal peptide of 38 amino acids but has no C-terminal extension. Amino acid sequence comparisons revealed that PpGns1 is 69% and 67% identical to citrus and soybean beta-1,3-glucanases, respectively. Southern analysis of total genomic DNA also indicates that at least three genes for beta-1,3-glucanases exist in peach, forming a small gene family. Characterization of four additional clones by PCR has identified a second beta-1,3-glucanase gene, PpGns2. PpGns2 has been partially sequenced, and when compared to PpGns1, it shows high sequence homology, 96% and 99% nucleotide identity in the first and (partial) second exons, respectively. The deduced partial sequence of the PpGns2 product displays only two differences from PpGns1 in the signal peptide and one in the (partial) mature protein (141 amino acids). The 5'-flanking promoter regions of these two genes share 90% identity in nucleotide sequences interrupted by five major gaps (4-109 nt long). The promoter region contains various sequences similar to cis-regulatory elements present in different stress-induced plant genes. In leaves and stems of peach shoot cultures grown in vitro, PpGns1 is induced within 12 h after exposure to a culture filtrate of Xanthomonas campestris pv. pruni or ethephon. However, it is not induced following treatment with mercuric chloride.