Toward an Understanding of the Molecular Mechanisms of Barnacle Larval Settlement: A Comparative Transcriptomic Approach

Abstract

BackgroundThe barnacle Balanus amphitrite is a globally distributed biofouler and a model species in intertidal ecology and larval settlement studies. However, a lack of genomic information has hindered the comprehensive elucidation of the molecular mechanisms coordinating its larval settlement. The pyrosequencing-based transcriptomic approach is thought to be useful to identify key molecular changes during larval settlement.Methodology and Principal FindingsUsing 454 pyrosequencing, we collected totally 630,845 reads including 215,308 from the larval stages and 415,537 from the adults; 23,451 contigs were generated while 77,785 remained as singletons. We annotated 31,720 of the 92,322 predicted open reading frames, which matched hits in the NCBI NR database, and identified 7,954 putative genes that were differentially expressed between the larval and adult stages. Of these, several genes were further characterized with quantitative real-time PCR and in situ hybridization, revealing some key findings: 1) vitellogenin was uniquely expressed in late nauplius stage, suggesting it may be an energy source for the subsequent non-feeding cyprid stage; 2) the locations of mannose receptors suggested they may be involved in the sensory system of cyprids; 3) 20 kDa-cement protein homologues were expressed in the cyprid cement gland and probably function during attachment; and 4) receptor tyrosine kinases were expressed higher in cyprid stage and may be involved in signal perception during larval settlement.ConclusionsOur results provide not only the basis of several new hypotheses about gene functions during larval settlement, but also the availability of this large transcriptome dataset in B. amphitrite for further exploration of larval settlement and developmental pathways in this important marine species.

The regulatory role of the NO/cGMP signal transduction cascade during larval attachment and metamorphosis of the barnacleBalanus (=Amphibalanus) amphitrite

“…To investigate whether the expression of NO/cGMP-related genes changes during the process of settlement, quantitative real-time PCR (qRT-PCR) of two key components of this signaling pathway, namely NOS and soluble guanylyl cyclase (sGC), were performed following protocols described elsewhere (Chen et al, 2011) using the NCBI Primer-BLAST program (Table2). Real-time PCR was carried out using a Kapa SYBR FAST qPCR master mix (Kapa Biosystems, Boston, MA, USA) and run on an ABI 7500 fast real-time PCR system (Applied Biosystems).…”

“…To test whether NO signaling may be involved in barnacle settlement, we took advantage of a high-throughput B. amphitrite transcriptome database generated in our laboratory (Chen et al, 2011). Our searches against the annotated transcriptome sequences revealed the presence of all major components of the NO signaling and related pathways, which include nitric oxide synthase (NOS), GC, phosphodiesterase (PDE) and cGMP-dependent protein kinase (PKG) (Table1; supplementary material TableS1, Fig.S1), suggesting that further investigation of this pathway in barnacle settlement was warranted.…”

The environmentally tuned transcriptomes ofMytilusmussels

“…osmotic stress, heat and high light) in the red seaweed Chondrus crispus and their results were among the first to show the prevalence of oxidative stress in intertidal organisms. In addition to describing physiological responses to abiotic factors like temperature and pH, transcriptomics has revealed the molecular mechanisms that underlie responses to predation in intertidal snails (Chu et al, 2014) and larval settlement in barnacles (Chen et al, 2011).…”

“Crustacea”: Cirripedia

“…A recent study on gene expression in nauplii II, VI, cyprids, and adults of the intertidal barnacle Amphibalanus amphitrite revealed a change in gene expression of certain genes during larval development (Chen et al 2011 ). Vitellogenin was only expressed in nauplius VI, suggesting it may be an energy source for the subsequent nonfeeding cypris stage.…”