Many marine invertebrates have planktonic larval and benthic juvenile/adult stages. When the planktonic larvae are fully developed, they must find a favorable site to settle and metamorphose into benthic juveniles. This transition from a planktonic to a benthic mode of life is a complex behavioral process involving substrate searching and exploration. Although the mechanosensitive receptor in the tactile sensor has been implicated in sensing and responding to surfaces of the substrates, few have been unambiguously identified. Recently, we identified that the mechanosensitive transient receptor potential melastatin-subfamily member 7 (TRPM7) channel, highly expressed in the larval foot of the mussel Mytilospsis sallei, was involved in substrate exploration for settlement. Here, we show that the TRPM7-mediated Ca²⁺ signal was involved in triggering the larval settlement of M. sallei through the calmodulin-dependent protein kinase kinase β/AMP-activated protein kinase/silk gland factor 1 (CaMKKβ-AMPK-SGF1) pathway. It was found that M. sallei larvae preferred the stiff surfaces for settlement, on which TRPM7, CaMKKβ, AMPK, and SGF1 were highly expressed. These findings will help us to better understand the molecular mechanisms of larval settlement in marine invertebrates, and will provide insights into the potential targets for developing environmentally friendly antifouling coatings for fouling organisms.

Marine biofouling caused by barnacle gregarious settlement poses significant challenges to various industries and ecosystems, such as increased drag on ship hulls, elevated fuel consumption, and heightened maintenance costs. While natural chemical cues are instrumental in driving barnacle settlement, the underlying mechanisms remain incompletely understood. In this work, we investigated the effects of adenosine (Ado), a settlement pheromone of Amphibalanus amphitrite cyprids, on cyprid exploration behavior, nano-mechanical properties of footprints, and gene expression using atomic force microscopy (AFM) and omics analysis. Results indicate that Ado significantly increases the settlement rate and exploration frequency of cyprids, and enhances the expression of the settlement-inducing protein complex (SIPC, which attracts other cyprids to settle in a gregarious manner). AFM results reveal that Ado-treated cyprids exhibit enhanced adhesion, self-healing, elasticity, and mechanical strength in their footprints, which may help them resist the shear forces from seawater. Transcriptome analysis suggests that Ado triggers the up-regulation of the transcription factors FTZ-F1 and Hr39, which may activate the 20E hormonal signaling pathway and promote the settlement process. Furthermore, Ado up-regulates the cement protein genes of CP19K-like4 and CP100K, which are involved in the initial adhesion process. These findings provide valuable insights into the role of pheromones in promoting barnacle settlement and offer a deeper understanding of the mechanisms driving this behavior.

Most fouling organisms have planktonic larval and benthic adult stages. Larval settlement, the planktonic–benthic transition, is the critical point when biofouling begins. However, our understanding of the molecular mechanisms of larval settlement is limited. In our previous studies, we identified that the AMP-activated protein kinase-silk gland factor 1 (AMPK-SGF1) pathway was involved in triggering the larval settlement in the fouling mussel M. sallei. In this study, to further confirm the pivotal role of SGF1, multiple targeted binding compounds of SGF1 were obtained using high-throughput virtual screening. It was found that the targeted binding compounds, such as NAD⁺ and atorvastatin, could significantly induce and inhibit the larval settlement, respectively. Furthermore, the qRT-PCR showed that the expression of the foot proteins’ genes was significantly increased after the exposure to 10 μM NAD⁺, while the gene expression was significantly suppressed after the exposure to 10 μM atorvastatin. Additionally, the production of the byssus threads of the adults was significantly increased after the exposure to 10–20 μM of NAD⁺, while the production of the byssus threads was significantly decreased after the exposure to 10–50 μM of atorvastatin. This work will deepen our understanding of SGF1 in triggering the larval settlement in mussels and will provide insights into the potential targets for developing novel antifouling agents.