Physiological characterization of the emergence from diapause: A transcriptomics approach

Abstract

Organisms inhabiting high-latitude environments have evolved adaptations, such as diapause to time reproduction and growth to optimize their survival. However, the physiological regulation of the timing of complex life histories is poorly understood, particularly for marine copepods, that diapause at depth. A member of the pelagic community of the sub-Arctic Pacific Ocean, Neocalanus flemingeri enters diapause in June. Egg production occurs in winter/spring. In order to characterize the transition from diapause to egg release, females were collected in late September from 400–700 m depth, incubated in the dark at 4–5 °C and sampled for RNASeq at weekly intervals. The diapause phenotype showed down-regulation of protein turnover and up-regulation of stress genes. Activation of the reproductive program was marked by the up-regulation of genes involved in germline development. Thereafter, progress through phases of oocyte development could be linked to changes in gene expression. At 5 weeks, females showed up-regulation of spermatogenesis, indicating that stored sperm had been in a quiescent stage and completed their maturation inside the female. Gene expression profiles provide a framework to stage field-collected females. The 7-week progression from diapause to late oogenesis suggests that females typically spawning in January initiated the reproductive program in November.

Capital Breeding in a Diapausing Copepod: A Transcriptomics Analysis

“…1,000 differentially expressed genes involved in reproduction, the progression through oogenesis is nearly synchronous in females collected in September from deep water (400 to 700 m) and incubated in the laboratory at 5 • C. The reproductive program starts with the up-regulation of genes involved in germline development, the first stage of oogenesis (OS0), and it takes 6-7 weeks before females start to spawn. While in most calanoid copepods all five stages of oogenesis are concurrent, Roncalli et al (2018b) found oogenesis to be sequential in N. flemingeri with processes associated with early oogenesis becoming down-regulated as females neared spawning.…”

“…The time series included weekly or semi-weekly sampling of deep-collected females followed by laboratory incubations starting at collection (Wk0) to 9.5 weeks post-collection (Wk9.5). In a previous study, the data from Wk0 to Wk7 were analyzed to characterize the reproductive program (Roncalli et al, 2018b).…”

“…As described previously (Roncalli et al, 2018b), Neocalanus flemingeri adult females were collected from depth on September 20, 2015 in Prince William Sound (station "PWS2"; depth:∼740 m; Latitude 60 • 32.1 N; Longitude 147 • 48.2 W) during the fall cruise of the Seward Line Long-term Observation Program (LTOP) 1 . Briefly, a multiple opening and closing plankton net (0.25 m 2 cross-sectional area; 150 µm mesh nets; Multinet-Midi, Hydro-Bios) was towed vertically from 700 m. Collections from 400-700 m were immediately diluted, and adult females of N. flemingeri were live sorted and either preserved within 35 min of the retrieval of the net ["Wk0", n = 6; RNAlater Stabilization Reagent (QIAGEN)] or sorted into 20L carboys for the incubation experiment.…”

“…These organisms typically resume feeding post-diapause -an effective bet hedging strategy that allows them to acquire resources for reproduction at two time points that are separated by the diapause period (Niehoff et al, 2002). However, the females of the sub-arctic copepod, Neocalanus flemingeri (Miller, 1988) rely on "capital" acquired during pre-diapause to fuel the entire reproductive program starting with early cell line differentiation ("germline formation"; Roncalli et al, 2018b). The annual life cycle of N. flemingeri is limited to a single generation per year with the early larval stages (nauplii) appearing in the water column during the winter (Miller and Clemons, 1988;Lenz and Roncalli, 2019).…”

“…In a recent study, the progression of oogenesis in N. flemingeri females was characterized using transcriptomics from diapause up to spawning (Roncalli et al, 2018b). Gene expression patterns in N. flemingeri were compared with the wellcharacterized reproductive program of the model species Drosophila melanogaster (Baker and Russell, 2009;Jagut et al, 2013;Kronja et al, 2014), followed by an analysis that linked gene expression in N. flemingeri to the five stages of oogenesis (OS0 -OS4) described for calanoid copepods using histology (Niehoff and Hirche, 1996;Niehoff, 2007).…”