Francisco M. Pinto

Mitochondrial DNA analysis of Northwest African populations reveals genetic exchanges with European, Near-Eastern, and sub-Saharan populations

Genetic studies have emphasized the contrast between North African and sub‐Saharan populations, but the particular affinities of the North African mtDNA pool to that of Europe, the Near East, and sub‐Saharan Africa have not previously been investigated. We have analysed 268 mtDNA control‐region sequences from various Northwest African populations including several Senegalese groups and compared these with the mtDNA database. We have identified a few mitochondrial motifs that are geographically specific and likely predate the distribution and diversification of modern language families in North and West Africa. A certain mtDNA motif (16172C, 16219G), previously found in Algerian Berbers at high frequency, is apparently omnipresent in Northwest Africa and may reflect regional continuity of more than 20000 years. The majority of the maternal ancestors of the Berbers must have come from Europe and the Near East since the Neolithic. The Mauritanians and West‐Saharans, in contrast, bear substantial though not dominant mtDNA affinity with sub‐Saharans.

Phylogeographic patterns of mtDNA reflecting the colonization of the Canary Islands

Although the Canary Islands were settled by humans, possibly of Berber origin, as late as 2500 years ago, the precise course and numbers of early migrations to the archipelago remain controversial. We have therefore analysed mtDNA variation (HVS‐I as well as selected RFLP sites) in 300 individuals from the seven Canary Islands. The distribution and variation across the islands in a specific mtDNA clade of Northwest African ancestry suggest that there was one dominant initial settlement process that affected all the islands, from east to west. This indicates that a certain genetic affinity of present‐day Canary Islanders to Northwest African Berbers mainly stems from the autochthonous population rather than slaves captured on the neighbouring African coast. The slave trade after the European conquest left measurable, though minor, traces in the mtDNA pool of the Canary Islands, which in its majority testifies to the European immigration.

Genetic relationship between the Canary Islanders and their African and Spanish ancestors inferred from mitochondrial DNA sequences

Nucleotide sequences of the hypervariable segment I of the control region of the mtDNA were determined in 101 individuals: 54 Canary Islanders, 18 North African Berbers, 18 Spanish mainlanders and 11 sub‐Saharan Guineans. In spite of the fact that only members of the Fang tribe were analysed, nucleotide diversity in Guineans (θ× 100 = 2·33)is one of the highest found in African populations.

Structure and evolution of the mitochondrial DNA complete control region in the Drosophila subobscura subgroup

The complete A + T‐rich region of mitochondrial DNA (mtDNA) has been cloned and sequenced in the species of the Drosophila subobscura subgroup D. subobscura, D. madeirensis and D. guanche. Comparative analysis of these sequences with others already published has identified new sequence motifs that are conserved in Drosophila and other insects. A putative bi‐directional promoter and a stop signal are proposed to be involved in the primary mtDNA strand replication of Drosophila. This region strongly resolves relationships of the species included in a phylogenetic analysis, both for closely related species and also at deeper phylogenetic levels when only the left and central domains are taken into account.

Characterization of tachykinin receptors in the uterus of the oestrogen-primed rat

The aim of our study was to characterize the tachykinin receptor population in the oestrogen‐primed rat uterus. For this purpose, we investigated the receptor type(s) responsible for tachykinin‐induced contraction of longitudinally‐arranged smooth muscle layer. The effects of substance P (SP), neurokinin A (NKA), neurokinin B (NKB) and several of their analogues with well‐defined selectivities for tachykinin NK1, NK2 and NK3 receptors were studied and their inhibition by the selective nonpeptide tachykinin receptor antagonists (S)1‐(2‐[3‐(3,4‐dichlorophenyl)‐1‐(3‐isopropoxyphenylacetyl)piperidin‐3‐yl]ethyl)‐4‐phenyl‐1‐azoniabicyclo[2.2.2]octane chloride (SR 140333, NK1‐selective), (S)‐N‐methyl‐N[4‐(4‐acetylamino‐4‐phenylpiperidino)‐2‐(3,4‐dichlorophenyl)butyl]benzamide (SR 48968, NK2‐selective) and (R)‐(N)‐(1‐(3‐(1‐benzoyl‐3‐(3,4‐dichlorophenyl)piperidin‐3‐yl)propyl)‐u20034‐phenylpiperidin‐u20034‐yl)‐N‐u2003methylacetamide (SR 142801, NK3‐selective) was evaluated. Additionally, expression of tachykinin receptor mRNA was examined by using the reverse transcription‐polymerase chain reaction (RT‐PCR). SP, NKA, [Nle10]‐NKA(4‐10), the analogue with selectivity at the tachykinin NK2 receptor type, and NKB elicited concentration‐dependent contractions of the rat uterus. The pD2 values were 5.95±0.19; 6.73±0.21; 7.53±0.12 and 5.76±0.21, respectively. The selective agonist for the tachykinin NK1 receptor [Sar9Met(O2)11]‐SP produced a small phasic response in the nanomolar concentration range. The selective tachykinin NK3 receptor agonist [MePhe7]‐NKB failed to induce any significant contraction. In the presence of the neutral endopeptidase inhibitor phosphoramidon (1u2003μM), the log concentration‐response curves to exogenous tachykinins and their analogues were shifted significantly leftwards. The pD2 values were 6.12±0.10, 8.04±0.07, 7.89±0.03 and 6.59±0.07 for SP, NKA, [Nle10]‐NKA(4‐10) and NKB, respectively. In the presence of phosphoramidon (1u2003μM), [Sar9Met(O2)11]‐SP (1u2003nM–0.3u2003μM) induced concentration‐dependent contractions of increasing amplitude when only one concentration of drug was applied to each uterine strip and the pD2 value was 7.61±0.89. [MePhe7]‐NKB induced small, inconsistent contractions and, therefore, a pD2 value could not be calculated. In experiments performed in the presence of phosphoramidon (1u2003μM), SR 48968 (3u2003nM–0.1u2003μM) caused parallel and rightward shifts in the log concentration‐response curves of NKA. The calculated pKB value was 9.16±0.08 and the slope of the Schild regression was 1.28±0.24. SR 48968 (0.1u2003μM) also antagonized responses to SP with an apparent pKB value of 7.63±0.13. SR 48968 (0.1u2003μM) inhibited contractions elicited by NKB (1u2003nM–3u2003μM) and [Nle10]‐NKA(4–10) (0.1u2003nM–3u2003μM) but had no effect on the response evoked by [Sar9Met(O2)11]‐SP (0.1u2003μM). SR 140333 (0.1u2003μM) inhibited responses to SP with an apparent pKB value of 7.19±0.22. This compound did not significantly affect responses to NKA, [Nle10]‐NKA(4‐10) and NKB, but suppressed [Sar9Met(O2)11]‐SP (0.1u2003μM)‐induced contraction. SR 142801 (0.1u2003μM) had no effect on responses to natural tachykinins or their analogues. Total RNA was extracted from some of the uteri used in functional studies. RT‐PCR assays revealed single bands corresponding to the expected product sizes encoding cDNA for tachykinin NK1 (587 base pairs) and NK2 receptors (491 base pairs) (n=6 different animals). A very low abundance transcript corresponding to the 325 base pairs product expected for the tachykinin NK3 receptor was detected. The present data show that functionally active tachykinin NK1 and NK2 receptors are expressed in the oestrogen‐primed rat uterus. The NK2 receptor type seems to be the most important one involved in the contractile responses elicited by tachykinins. NK3 receptors are present in trace amounts and seem not to be involved in tachykinin‐induced contractions.

Human enzyme polymorphism in the Canary Islands. VI. Northwest African influence.

The genetic polymorphism of eight red cell enzymes was analyzed in population samples from the Northwest African Continent and from the South of Spain in order to study their genetic relationships with the Canarian population. The Moroccan, Berber and Spanish populations, although geographically more distant from the Canary Islands than the Saharan and Mauritanian ones, are genetically more closely related to the Canarian population. The glucose-6-phosphate dehydrogenase Gc allele earlier found only in the Canary Islands was detected in the Berber sample. The Spanish, Berber and African Black contributions to the Canarian hybrid population was estimated to 70, 20 and 10%, respectively.

Regulation by oestrogens of tachykinin NK3 receptor expression in the rat uterus

The expression of the tachykinin NK3 receptor and its regulation by ovarian steroids were analyzed by reverse transcription-polymerase chain reaction (RT-PCR) in uteri from ovariectomized rats. A single transcript corresponding to the 325-bp product expected for the tachykinin NK3 receptor was detected in uteri from olive oil-treated (control) ovariectomized rats. The level of tachykinin NK3 receptor mRNA in progesterone-treated animals was similar to that observed in uteri from control ones. Tachykinin NK3 receptor mRNA levels were significantly smaller in uteri from oestrogen-treated ovariectomized rats, with approximately a 32-fold decrease. These findings suggest that oestrogen, but not progesterone, regulates the expression of tachykinin NK3 receptors in the rat uterus.

Molecular diagnosis of alkaptonuria mutation by analysis of homogentisate 1,2 dioxygenase mRNA from urine and blood

Alkaptonuria (AKU) is caused by lack of homogentisate 1, 2 dioxygenase (HGO) activity. From the complete sequence of a human HGO cDNA, primers were designed in order to obtain reverse transcription-polymerase chain reaction products from tissues with ectopic transcription amenable to diagnostic analysis. A search for mutations in HGO cDNA was performed in an AKU family using urine and blood samples. The results show complete cosegregation (Z = 6.32; theta = 0) between a C-->T transition at position 817 of the human HGO cDNA and AKU. This mutation predicts a Pro-->Ser replacement at amino acid 230, and generates an EcoRV site.

Molecular genetic characterization of plant somatic hybrids

An efficient and easy method for genetic characterization of plant somatic hybrids is proposed. In a first qualitative approach, four somatic hybrids and their parental species (Nicotiana tabacum andN. plumbaginifolia) were characterized by DNA fingerprinting and Random Amplification of Polymorphic DNA (RAPD). After this, a quantitative estimation of the degree of parental contribution to the hybrids was carried out by means of a slot-blot analysis. Both qualitative methods, showed one hybrid identical toN. tabacum, two almost identical toN. plumbaginifolia, and a fourth similar to this parental species, but with someN. tabacum admixture. The quantitative method, for the same hybrids, gave 83%, 7%, 7%, and 37%N. tabacum DNA contribution, respectively.

TRANSFER OF KANAMYCIN RESISTANCE FROM NICOTIANA TABACUM TO NICOTIANA PLUMBAGINIFOLIA BY FUSION OF X-IRRADIATED PROTOPLASTS

SummaryNuclear hybrids have been obtained by fusion of mesophyll protoplasts ofNicotiana plumbaginifolia and x-irradiated or iodoacetate-treated mesophyll protoplasts of a kanamycin-resistant line ofN. tabacum. The effect of irradiation on the recovery of asymmetric hybrids was evaluated by analysis of their morphology, fertility, chromosome number, isozyme patterns, restriction patterns in their organelle DNAs, and presence of the kanamycin-resistance gene. The results presented in this paper show that x-ray irradiation leads to a significant reduction in the amount ofN. tabacum genome present in the hybrids and demonstrates, once more, the power of this technique to induce directional loss of genomic traits of the irradiated parent.