J Mol Evol (1991) 33:83-91

Journal of Molecular Evolution (~) Springer-Verlag New York Inc. 1991

The Main Regulatory Region of Mammalian Mitochondrial DNA: Structure-Function Model and Evolutionary Pattern Cecilia Saccone, ~ Graziano Pesole, ~ and Elisabetta Sbis~ 2 Dipartimento di Biochimicae BiologiaMoleeolare,Universitfidi Bari, Italy 2Centro di Studio sui Mitocondrie MetabolismoEnergetieo,CNR, Bail, Italy

Summary.

The evolution o f the main regulatory region (D-loop) o f the mammalian mitochondrial genome was analyzed by comparing the sequences o f eight mammalian species: human, c o m m o n chimpanzee, pygmy chimpanzee, dolphin, cow, rat, mouse, and rabbit. The best alignment o f the sequences was obtained by optimization of the sequence similarities c o m m o n to all these species. The two peripheral left and right D-loop domains, which contain the main regulatory elements so far discovered, evolved rapidly in a species-specific manner generating heterogeneity in both length and base composition. They are prone to the insertion and deletion o f elements and to the generation o f short repeats by replication slippage. However, the preservation o f some sequence blocks and similar cloverleaf-like structures in these regions, indicates a basic similarity in the regulatory mechanisms o f the m i t o c h o n d r i a l g e n o m e in all mammalian species. We found, particularly in the right domain, significant similarities to the telomeric sequences of the mitochondrial (mt) and nuclear D N A of Tetrahymena thermophila. These sequences may be interpreted as relics o f telomeres present in ancestral linear forms o f m t D N A or may simply represent efficient templates o f R N A primase-like enzymes. Due to their peculiar evolution, the two peripheral domains cannot be used to estimate in a quantitative way the genetic distances between mammalian species. On the other hand the central domain, highly conserved during evolution, behaves as a good molecular clock. Reliable estimates o f the times o f divergence between closely and distantly related species were ob-

Offprint requests to: C. Saccone

tained from the central domain using a Markov model and assuming nonhomogeneous evolution o f nucleotide sites.

Key words: Mammalian mitochondrial DNA -Origin of replication -- Mitochondrial DNA evolution -- Stationary Markov model -- Phylogenetic tree -- Telomeres -- D-loop -- Regulatory region

Introduction The presence of only one major noncoding segment in the mitochondrial genome is a feature c o m m o n to all metazoa. In vertebrates this region, spanning between the Phe- and Pro-tRNA genes, is called the D-loop-containing region because of the threestranded displacement (D) loop structure created by the nascent heavy (H) strand at the level o f the Hstrand replication origin (On). It also contains promoters for the transcription o f both the heavy Strand (HSP) and the light strand (LSP). This region is the target site for numerous proteins and enzymes, such as D N A and RN A polymerases and transcription and regulatory factors and is thus subjected t o various evolutionary pressures. Because all these proteins are coded for by nuclear DNA, the study o f the D-loop-containing region is also extremely important for shedding light on the processes inherent in nucleus-mitochondrion coevolution. In order to gain deeper insight into the evolutionary dynamics o f the noncoding region o f the mammalian mitochondrial genome, we undertook a detailed investigation of its evolution at the molecular level. In previous papers we have identified several well-preserved features in the evolution of

84

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20 30 40 50 60 70 80 90 I00 T TCT TTCATGGGGAAGCAAATTTAAGTGCCACCCAAGTATTGG ............................................................. :::::::::::::::::::::::: :: ::::: ::::: ::: TTCT TT CATC~C-GCAAGCAAATT TAG~TACCACC TAAGTACT GG .............................................................

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150 160 170 180 190 200 210 220 230 240 TAAC-CGCTATGTATT-TCGTAcATTACTG-CCA--GCCACCATGAATA--TTACATAGTACTATAATCATTTAACCACCTATAACACATAAA :11 :::11111:1: I:ll:lll::::: ::: ::::111::::: : : :11:: :1: ::: ::: ::::::: ::::::: CAA•-CC-CTATGTATT-TCGTA•ATTA•TG-CCA--G••AC•ATGAATA--TCGTA•AGTACCATA-TCACCCAACTACCTATAGTACATAAA ::11 :::11111:1: I:11:111::::: ::: ::::111::::: : :::: I1::111:. 11 :1 1::: ::::;:::::: CAAC-•GCTATGTATT-TCGTACATTACTG-CCA•-GCCA•CATGAATA--TTGTACGGTACCATAAATACTTGACCACCT•TAGTACATAAA II 11111111:: I II III: : : III: II I::: : : : :: : :::::: T•C•T•TCTTTGTAAATATT•ATATA••TACATCCTATGTATTATTGTGCATTCATTT--ATTT---CCATACG-A-TAA ..... GT .... TAAAG-CCCGTATTAAT-TA-T-CATTAA : I : : I ::: : I1:: Illit:l :l II 111: : :: :: : - II1: : 9 :::. 9 I1...I : : :" TAC-ATAACATTA-AT-GTA-ATAAA--GA~ATAATATGTAT-ATAGTA~ATTAAATT--ATATGCCCCATGCATA-TAAGCAA~GTA~ATGACC~TCTATAG ................ ::1 ::11 I: 1 1: : :.:: 11 II11 II1:111:1 I:1111:::1:: 1: : 1:111:111 11:::1 I1: :1 "::1 9 . TAC-ATAAAATGATATGG-ACATTAA--AACATT-TATGTAT-ATCGTACATTAAATT--ATTTTCCCCAAGCATA-TAAGCAT-GTA--ATATATATCTAATGATTT ............ ::1 ::::: : : :: :-::: : II::: Illll:l :l;l{:ill:::: : :::::::ill::::: ;:::: II:. :+ : ::: :::: :: TAC-ATAAATTTACATAGTACAACAG--TACATT-TATGTAT•ATCGTACATTAAACT--ATTTTCCCCAAGCATA-TAACCTA-GTAC•ATTAA-ATC-AATGGTT• ............ :1 : : : : : : : :::: :::;; II;; IIIII I :l:ll III ::: :.. : ::111 : :: ::::::: If:: :1:: 9 9 :: : : . . . . . . AACAATAAAT-T-CATAA-CCAACATTTAACATACTATGTTTAATcGTGCAT-AAATTCCTCATCCCCCATGAATAATAAGCTA-GTAC-ATTACTGCTTGATTGGACATAATCCACT--

250 260 270 280 290 300 310 320 330 340 350 360 _CAGTACATAGCACATACAATTATATACCGTACATAGCACATTACAGTCAAATCCATCCTCGCCCCCACGGATG .............................................. ::: :11::: I:1::::: ::: :::::::::::::::::::::::::I 1: :::1:::::::::::I:: _CAGAACATAGTACATACAACCATACACCGTACATAGCACATTACAGTCAAACCCCTCCTCCCCCCCACGGATG .............................................. ::: :11::::::1:: : :::: :::::::::::::::::::::::::: :::1 :1:: ::::: ::::: CAGTACATACTACATAAAGCCATTTACCGTACATAGCACATTACACTCAAATCCCTTCTCGTCCCCATGGATG .............................................. ::11:: :::1: : : I : :1:11: I :: : I: 1: I : :: : •TTTTACATATTACATGATATGTATAATCTTACATATTATATATCCCCTAACAATTTTATTTCCATTATACCTATGGTCGCT ...... CCATTAGATCACGAG ................. ::ll:: :1:I: :: :: I :1:11::1 :: I1 I1 1: : :::: :::: : : ::::::::::::::: ~CAGTACATAATAC~TATAATTATTGACTGTACATAGTACATT-ATCTCAAAT~CATTCTTGATAGTATATCTATTATATATTCCTTACCATTAGATcACGAG .................

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