ABSTRACTNitrogen is one of the major nutrients limiting microbial productivity in the ocean, and as a result, most marine microorganisms have evolved systems for responding to nitrogen stress. The highly abundant alphaproteobacterium “CandidatusPelagibacter ubique,” a cultured member of the orderPelagibacterales(SAR11), lacks the canonical GlnB, GlnD, GlnK, and NtrB/NtrC genes for regulating nitrogen assimilation, raising questions about how these organisms respond to nitrogen limitation. A survey of 266Alphaproteobacteriagenomes found these five regulatory genes nearly universally conserved, absent only in intracellular parasites and members of the orderPelagibacterales, including “Ca. Pelagibacter ubique.” Global differences in mRNA and protein expression between nitrogen-limited and nitrogen-replete cultures were measured to identify nitrogen stress responses in “Ca.Pelagibacter ubique” strain HTCC1062. Transporters for ammonium (AmtB), taurine (TauA), amino acids (YhdW), and opines (OccT) were all elevated in nitrogen-limited cells, indicating that they devote increased resources to the assimilation of nitrogenous organic compounds. Enzymes for assimilating amine into glutamine (GlnA), glutamate (GltBD), and glycine (AspC) were similarly upregulated. Differential regulation of the transcriptional regulator NtrX in the two-component signaling system NtrY/NtrX was also observed, implicating it in control of the nitrogen starvation response. Comparisons of the transcriptome and proteome supported previous observations of uncoupling between transcription and translation in nutrient-deprived “Ca.Pelagibacter ubique” cells. Overall, these data reveal a streamlined, PII-independent response to nitrogen stress in “Ca.Pelagibacter ubique,” and likely otherPelagibacterales, and show that they respond to nitrogen stress by allocating more resources to the assimilation of nitrogen-rich organic compounds.IMPORTANCEPelagibacteralesare extraordinarily abundant and play a pivotal role in marine geochemical cycles, as one of the major recyclers of labile dissolved organic matter. They are also models for understanding how streamlining selection can reshape chemoheterotroph metabolism. Streamlining and its broad importance to environmental microbiology are emerging slowly from studies that reveal the complete genomes of uncultured organisms. Here, we report another remarkable example of streamlined metabolism inPelagibacterales, this time in systems that control nitrogen assimilation.Pelagibacteralesare major contributors to metatranscriptomes and metaproteomes from ocean systems, where patterns of gene expression are used to gain insight into ocean conditions and geochemical cycles. The data presented here supply background that is essential to interpreting data from field studies.