Marine Biotoxins Program, NOAA National Ocean Service, Center for Coastal Environmental Health and Biomolecular Research, Charleston, SC 29412, USA.
BMC Genomics. 2011 Jul 5;12:346. doi: 10.1186/1471-2164-12-346.
The role of coastal nutrient sources in the persistence of Karenia brevis red tides in coastal waters of Florida is a contentious issue that warrants investigation into the regulation of nutrient responses in this dinoflagellate. In other phytoplankton studied, nutrient status is reflected by the expression levels of N- and P-responsive gene transcripts. In dinoflagellates, however, many processes are regulated post-transcriptionally. All nuclear encoded gene transcripts studied to date possess a 5' trans-spliced leader (SL) sequence suggestive, based on the trypanosome model, of post-transcriptional regulation. The current study therefore sought to determine if the transcriptome of K. brevis is responsive to nitrogen and phosphorus and is informative of nutrient status.
Microarray analysis of N-depleted K. brevis cultures revealed an increase in the expression of transcripts involved in N-assimilation (nitrate and ammonium transporters, glutamine synthetases) relative to nutrient replete cells. In contrast, a transcriptional signal of P-starvation was not apparent despite evidence of P-starvation based on their rapid growth response to P-addition. To study transcriptome responses to nutrient addition, the limiting nutrient was added to depleted cells and changes in global gene expression were assessed over the first 48 hours following nutrient addition. Both N- and P-addition resulted in significant changes in approximately 4% of genes on the microarray, using a significance cutoff of 1.7-fold and p ≤ 10-4. By far, the earliest responding genes were dominated in both nutrient treatments by pentatricopeptide repeat (PPR) proteins, which increased in expression up to 3-fold by 1 h following nutrient addition. PPR proteins are nuclear encoded proteins involved in chloroplast and mitochondria RNA processing. Correspondingly, other functions enriched in response to both nutrients were photosystem and ribosomal genes.
Microarray analysis provided transcriptomic evidence for N- but not P-limitation in K. brevis. Transcriptomic responses to the addition of either N or P suggest a concerted program leading to the reactivation of chloroplast functions. Even the earliest responding PPR protein transcripts possess a 5' SL sequence that suggests post-transcriptional control. Given the current state of knowledge of dinoflagellate gene regulation, it is currently unclear how these rapid changes in such transcript levels are achieved.
在佛罗里达州沿海水域，沿海营养源在保持膝沟藻赤潮方面的作用是一个有争议的问题，这需要研究这种甲藻中营养响应的调节。在其他研究过的浮游植物中，营养状况反映在 N 和 P 反应性基因转录本的表达水平上。然而，在甲藻中，许多过程是转录后调节的。迄今为止，所有研究过的核编码基因转录本都具有 5' 反式拼接前导（SL）序列，根据锥虫模型，提示转录后调节。因此，本研究旨在确定膝沟藻的转录组是否对氮和磷有反应，并能提供营养状况的信息。
对 N 耗尽的膝沟藻培养物进行的微阵列分析显示，与营养充足的细胞相比，参与 N 同化（硝酸盐和铵转运体、谷氨酰胺合成酶）的转录本表达增加。相比之下，尽管基于其对 P 添加的快速生长反应有 P 饥饿的证据，但 P 饥饿的转录信号并不明显。为了研究营养添加对转录组的反应，将限制营养物添加到耗尽的细胞中，并在添加营养物后的头 48 小时内评估全局基因表达的变化。氮和磷的添加都导致微阵列上大约 4%的基因发生显著变化，使用 1.7 倍的显著性截止值和 p ≤ 10-4。到目前为止，在两种营养处理中，最早响应的基因都以五肽重复（PPR）蛋白为主，这些蛋白在营养添加后 1 小时内表达增加了 3 倍。PPR 蛋白是参与叶绿体和线粒体 RNA 加工的核编码蛋白。相应地，其他对两种营养物质都有反应的功能是光合作用系统和核糖体基因。
微阵列分析为膝沟藻中的 N 限制而非 P 限制提供了转录组证据。对添加 N 或 P 的转录组反应表明，存在一个协同的程序，导致叶绿体功能的重新激活。即使是最早响应的 PPR 蛋白转录本也具有 5' SL 序列，提示转录后控制。考虑到目前对甲藻基因调控的了解程度，目前尚不清楚如何实现这些转录水平的快速变化。
