It is thought that during latent infection, Mycobacterium tuberculosis bacilli 
are retained within granulomas in a low-oxygen environment. The dormancy 
survival (Dos) regulon, regulated by the response regulator DosR, appears to be 
essential for hypoxic survival in M. tuberculosis, but it is not known how the 
regulon promotes survival. Here we report that mycobacteria, in contrast to 
enteric bacteria, do not form higher-order structures (e.g. ribosomal dimers) 
upon entry into stasis. Instead, ribosomes are stabilized in the associated form 
(70S). Using a strategy incorporating microfluidic, proteomic, and ribosomal 
profiling techniques to elucidate the fate of mycobacterial ribosomes during 
hypoxic stasis, we show that the dormancy regulator DosR is required for optimal 
ribosome stabilization. We present evidence that the majority of this effect is 
mediated by the DosR-regulated protein MSMEG_3935 (a S30AE domain protein), 
which is associated with the ribosome under hypoxic conditions. A Δ3935 mutant 
phenocopies the ΔdosR mutant during hypoxia, and complementation of ΔdosR with 
the MSMEG_3935 gene leads to complete recovery of dosR mutant phenotypes during 
hypoxia. We suggest that this protein is named ribosome-associated factor under 
hypoxia (RafH) and that it is the major factor responsible for DosR-mediated 
hypoxic survival in mycobacteria.