Using anti-(U3)RNP autoantibodies, we have isolated and characterized two additional small nucleolar RNAs from HeLa cells, which are less abundant than U3 RNA. Both RNAs possess a trimethylguanosine cap as judged by precipitation with anti-TMG antibody, but are not precipitated by either anti-Sm or anti-La antibodies. In addition, both RNAs are not precipitable by anti-Th serum, which recognizes another nucleolar RNP autoantigen. Sequence analysis revealed that one of these RNAs, 136 nucleotides long, is the human U8 homolog; while the other, 105 nucleotides long, represents a novel species which we designate U13. Both RNAs share with U3 two conserved sequences (boxes C and D). The role of one or both of these boxes in binding the common 34 kd antigenic protein, otherwise known as fibrillarin, is discussed. Fractionation of HeLa cells revealed that U8 and U13, like U3, reside in the nucleolus. In glycerol gradients both RNAs cosediment with larger structures possibly representing ribosomal precursors. We propose that U3, U8 and U13 comprise a new subset of mammalian snRNPs whose roles in ribosome biogenesis are discussed.