Master of Philosophy (Medicine) / The actin cytoskeleton is important in many cellular processes such as motility, and establishing and maintaining cell morphology. Members of the tropomyosin protein family associate with the actin cytoskeleton along the major groove of actin filaments (F-actin), stabilising them and regulating actin-filament dynamics. To date over 40 non-muscle tropomyosin isoforms have been identified, which are encoded by 4 different genes (α, β, γ, δ). Individual tropomyosin isoforms define functionally distinct F-actin populations. Previous studies have shown that tropomyosins sort to distinct subcellular compartments at different stages of development in polarised cells. Neuronal growth cones are highly dynamic polarised structures, dependent on a constant reorganisation of the actin cytoskeleton. By eliminating tropomyosins in a knockout (KO) mouse model, we investigated the role of two tropomyosin isoforms, Tm5NM1 and Tm5NM2 (γTm gene products) in growth cone dynamics and neurite outgrowth. Growth cone protrusion rates were significantly increased in one day old Tm5NM1/2 KO hippocampal neurons compared to WT controls. Neuritogenesis was significantly affected by the elimination of Tm5NM1/2, with a slight decrease in neurite length and an increase in neuronal branching in neurons cultured for four days. At the molecular level, the depletion of Tm5NM1/2 had no impact on the protein levels and activity of ADF/cofilin in hippocampal neurons while in cortical neurons a subtle but significant increase in ADF/cofilin activity was observed. The subtle phenotype in the early stages of neuritogenesis observed from eliminating Tm5NM1/2 may be explained with functional compensation by other tropomyosin isoforms. Functional compensation for the loss of Tm5NM1/2 may be provided by isoforms Tm5a/5b, TmBr2 and Tm4 as they localise to the growth cones, structures where Tm5NM1/2 are normally found. These results suggest that Tm5NM1/2 may not be required for early stages of neuritogenesis but may still play a fine-tuning role for this process.

Master of Philosophy (Medicine) / The actin cytoskeleton is important in many cellular processes such as motility, and establishing and maintaining cell morphology. Members of the tropomyosin protein family associate with the actin cytoskeleton along the major groove of actin filaments (F-actin), stabilising them and regulating actin-filament dynamics. To date over 40 non-muscle tropomyosin isoforms have been identified, which are encoded by 4 different genes (α, β, γ, δ). Individual tropomyosin isoforms define functionally distinct F-actin populations. Previous studies have shown that tropomyosins sort to distinct subcellular compartments at different stages of development in polarised cells. Neuronal growth cones are highly dynamic polarised structures, dependent on a constant reorganisation of the actin cytoskeleton. By eliminating tropomyosins in a knockout (KO) mouse model, we investigated the role of two tropomyosin isoforms, Tm5NM1 and Tm5NM2 (γTm gene products) in growth cone dynamics and neurite outgrowth. Growth cone protrusion rates were significantly increased in one day old Tm5NM1/2 KO hippocampal neurons compared to WT controls. Neuritogenesis was significantly affected by the elimination of Tm5NM1/2, with a slight decrease in neurite length and an increase in neuronal branching in neurons cultured for four days. At the molecular level, the depletion of Tm5NM1/2 had no impact on the protein levels and activity of ADF/cofilin in hippocampal neurons while in cortical neurons a subtle but significant increase in ADF/cofilin activity was observed. The subtle phenotype in the early stages of neuritogenesis observed from eliminating Tm5NM1/2 may be explained with functional compensation by other tropomyosin isoforms. Functional compensation for the loss of Tm5NM1/2 may be provided by isoforms Tm5a/5b, TmBr2 and Tm4 as they localise to the growth cones, structures where Tm5NM1/2 are normally found. These results suggest that Tm5NM1/2 may not be required for early stages of neuritogenesis but may still play a fine-tuning role for this process.

Thesis (Ph. D.)--University of Washington, 2007. / Vita. Includes bibliographical references (leaves 152-168).

Diese Arbeit beschreibt die immunologischen Eigenschaften von Acanthochilonema viteae Tropomyosin, einem Muskel-assoziierten Protein. A. viteae ist ein zu den Filarien gehörender Parasit von Gerbilen, ähnlich dem humanpathogenen Filarie Onchocercha volvulus. Diese Arbeit hatte die funktionelle Charakterisierung von A. viteae Tropomyosin im Kontext der natürlichen Infektion und experimentellen Vakzinierung zum Ziel. Das allergene Potential des Tropomyosins und die Produktion von spezifischen IgE-Antikörpern wurden untersucht. Außerdem wurden Tropomyosin-spezifische monoklonale Antikörpern (mAk) entwicklet. Es konnte gezeigt werden, dass Tropomyosin als vielversprechender Kandidat zur Vakzinentwicklung gegen Filariosen angesehen werden kann, wobei berücksichtigt werden muss, dass deutliche Effekte nur unter Th1-Bedingungen auftreten. Die Vakzinierung mit Protein oder DNA reduzierte Adultwurmzahlen um 30 bzw. 45%. Während einer Infektion fungiert Tropomyosin als funktionelles Allergen und führt zur Produktion von hohen Mengen spezifischen IgEs. Ein Screening synthetischer Peptid-Bibliotheken zeigte gemeinsame 13 IgG- und 11 IgE-Epitope. Weiters konnte Kreuzreaktivität mit anderen Tropomyosinen und diesen gemeinsamen IgE-Epitopen nachgewiesen werden. Mit Hilfe von mAk konnte gezeigt werden, dass Tropomyosin auf der Oberfläche der Kutikula der Larvenstadien L3 und microfilariae des Parasiten vorkommt. Durch die Deglykosylierung des nativen Proteins wurde deutlich, dass einige Epitope von posttranslationellen Modifikationen gebildet werden. Weitere Immunisierungen mit Tropomyosin führten zu einem ähnlichen Profil der Zellaktivierung und Antikörperproduktion wie der Adjuvant Aluminiumhydroxid. Jedoch führte die Behandlung zur IL-10 Produktion und zur Zunahme von Gr1+/ CD11b+ Zellpopulationen, welche natürliche Surpressors darstellen. Zusammenfassend kann gesagt werden, dass A. viteae Tropomyosin immunmodulierende Eigenschaften aufweist und als Komponente eines zu entwickelnden Vakzins in Frage kommt. / This study describes the immunological properties of Acanthocheilonema viteae muscle-associated protein tropomyosin. A. viteae is a filarial parasite of jirds that resembles the important human parasite Onchocerca volvulus. Focus of experiments is on unraveling the functional properties of tropomyosin in the context of an infection and experimental vaccination. Additionally, allergenic potential of tropomyosin was investigated and the ability to induce high levels of specific IgE. A part of the study was also aimed at the development of anti-tropomyosin monoclonal antibodies (mAb). This study revealed that tropomyosin is a promising antigen for vaccines against filarial nematodes, however, effective only in a Th1 biased environment. Vaccination with protein or DNA resulted in 30% - 45% protection that was not associated with specific IgG or IgE. During infection tropomyosin is an allergen and leads to the production of high levels of specific IgE. Screening of synthetic peptide libraries showed 13 IgG and 11 IgE co-located epitopes and revealed cross-reactivity with other tropomyosins and sharing of IgE epitopes. mAb were raised against A. viteae tropomyosin and showed that tropomyosin is abundant on the cuticle of L3 and microfilariae of the parasite. Deglycosylation of the native protein showed that some epitopes were formed by the posttranslational modifications. Additionally, immunization shows that tropomyosin induces a similar pattern of cell activation and antibody production as aluminium hydroxide adjuvant, but leads to the induction of IL-10 and the increase of population of GR1+/CD11b+ cells. These cells are regarded as natural suppressors. Taken together, results show that A. viteae tropomyosin has immunomodulating properties and can be considered as a component of an efficient vaccine.

Yeung Kai-shing, Elvis. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2000. / Includes bibliographical references (leaves 98-104). / Abstracts in English and Chinese. / Abstract (English) --- p.i / Abstract (Chinese) --- p.iii / Acknowledgement --- p.v / Table of Contents --- p.vii / List of Tables --- p.x / List of Figures --- p.xi / Abbreviations --- p.xiii / Chapter Chapter 1 --- General Introduction --- p.1 / Chapter Chapter 2 --- Literature Review / Chapter 2.1 --- Allergy to shellfish --- p.3 / Chapter 2.2 --- Identification and characterization of crustacean allergens --- p.7 / Chapter 2.3 --- Identification and characterization of mollusk allergens --- p.10 / Chapter 2.4 --- Cross-reactivity between shellfish allergens --- p.14 / Chapter 2.5 --- Epitopes of shellfish allergens --- p.17 / Chapter Chapter 3 --- Isolation and characterization of allergens in cephalopods / Chapter 3.1 --- Introduction --- p.22 / Chapter 3.2 --- Materials and methods --- p.25 / Chapter 3.2.1 --- Animals --- p.25 / Chapter 3.2.2 --- Sera --- p.25 / Chapter 3.2.3 --- Cephalopod tissue extracts --- p.25 / Chapter 3.2.4 --- Quantitation of protein --- p.27 / Chapter 3.2.5 --- Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) --- p.27 / Chapter 3.2.6 --- Immunoblotting --- p.29 / Chapter 3.2.7 --- Immunological detection of IgE binding protein --- p.30 / Chapter 3.2.8 --- Controlling of ribonuclease (RNase) activity --- p.31 / Chapter 3.2.9 --- Isolation of total RNA --- p.31 / Chapter 3.2.10 --- Construction of cDNA library --- p.32 / Chapter 3.2.10.1 --- Synthesis of first-strand cDNA --- p.33 / Chapter 3.2.10.2 --- Amplification of cDNA by LD-PCR --- p.33 / Chapter 3.2.10.3 --- Polishing of ds cDNA --- p.34 / Chapter 3.2.10.4 --- Ligation of adaptor --- p.34 / Chapter 3.2.10.5 --- Phosphorylation of adaptor-ligated cDNA --- p.35 / Chapter 3.2.10.6 --- Size fractionation of cDNA --- p.35 / Chapter 3.2.10.7 --- Ligation of cDNA to vector --- p.36 / Chapter 3.2.10.8 --- Packaging of plasmids to bacteriophage --- p.36 / Chapter 3.2.10.9 --- Titration of the packaging reaction --- p.38 / Chapter 3.2.10.10 --- Amplification of the cDNA library --- p.39 / Chapter 3.2.11 --- Immunoscreening of octopus cDNA library --- p.40 / Chapter 3.2.12 --- Extraction of phage DNA --- p.42 / Chapter 3.2.13 --- Purification of cDNA insert from phage plasmid --- p.42 / Chapter 3.2.14 --- cDNA cloning and protein expression --- p.43 / Chapter 3.2.15 --- Nucleotide sequencing analysis of DNA --- p.45 / Chapter 3.2.16 --- Immunoblot analysis of the recombinant allergen --- p.46 / Chapter 3.3 --- Results / Chapter 3.3.1 --- Immunoblotting of allergens in cephalopod --- p.47 / Chapter 3.3.2 --- RNA from octopus --- p.47 / Chapter 3.3.3 --- The cDNA library of octopus --- p.51 / Chapter 3.3.4 --- Immunoscreening of cDNA library from octopus --- p.56 / Chapter 3.3.5 --- Nucleotide sequencing determination and analysis --- p.61 / Chapter 3.3.6 --- Protein expression of Oct l I --- p.76 / Chapter 3.3.7 --- Immunoblot analysis of recombinant tropomyosin from octopus --- p.83 / Chapter 3.4 --- Discussion --- p.87 / Chapter Chapter 4 --- Conclusions --- p.96 / References --- p.98

Die hypertrophe Kardiomyopathie ist eine autosomal dominant vererbte Herzmuskelerkrankung. Es kommt zu einer asymmetrischen Hypertrophie insbesondere des Herzmuskels. Symptome sind unspezifisch und reichen von Dyspnoe bis hin zu Synkopen. Gelegentlich ist der plötzliche Herztod die erste Manifestation der Erkrankung. Molekulargenetische Untersuchung des Genomes dieser Patienten zeigten, dass diese Patienten Mutationen im Proteinen des Sarkomers aufwiesen. Hierunter fällt das beta-MHC-Gen und alpha-Tropomysin-Gen. Wir untersuchten 45 nicht miteinanderverwandte Patienten auf Mutationen im beta-MHC-Gen und im alpha-Tropomysin-Gen. Folgende molekulargenetische Untersuchungen wurden angewendet: Polymerase-Ketten-Reaktion, Single-Strand-Polymorphismus-Anlyse und Sequenzierung. Bei 6 Patienten (13%) fanden wir eine Mutation im beta-Myosin-Gen. Kein Patient hatte eine Mutation im alpha-Tropomyosin-Gen. In der Literatur wird eine Mutation im beta-MHC-Gen in bis zu 35% und im alpha-Tropomyosin-Gen in bis zu 3% der Fälle angenommen. Unsere Patienten hatten die Mutationen: Exon 13 Arg403Trp und Val411Ile, Exon 19 Arg719Trp, Exon 20 Ile736Thr, Exon 21 Leu796Phe und Exon 23 Cys905Phe. Die Mutationen Arg403Trp und Arg719Trp waren vorher bereits bekannt. Die Mutationen, Ile736Thr, Val411Ile, Leu796Phe und Cys905Phe wurden in der Form von uns erstmals ermittelt. Offensichtlich besitzen unsere Patienten überdurchschnittlich häufiger Mutationen in anderen Genen, die in dieser Studie nicht untersucht wurden. Der Phänotyp der Krankheit der HCM war bei unserem Patientenkollektiv sehr heterogen, und es ließen sich keine signifikanten Unterscheidungen eststellen. So haben wir uns darauf beschränkt, bei den 6 Patienten mit Mutationen nur nach den von Burn et al. (1997) aufgestellten Risikofaktoren zu suchen, die einen plötzlichen Herztod herbeiführen können, und haben sie danach in Patienten mit einem höheren oder niedrigeren Risiko eingestuft. Kriterien für ein erhöhtes Risiko, einen plötzlichen Herztod zu erleiden, wurden von Patienten mit den Mutationen: Exon 13 Arg403Trp, Exon 13 Val411Ile, Exon 19 Arg719Trp, Exon 20 Ile736Thr und Exon 21 Leu796Phe erfüllt. Mutationen an diesen Positionen sind auch in der Literatur mit einem erhöhten Risiko für einen plötzlichen Herztod assoziiert worden. Der Patient mit der Mutation im Exon 23 Cys905Phe wies wenige Risikofaktoren auf und unterscheidet sich somit nicht von den in der Literatur beschriebenen Patienten. Wir konnten dies mit unseren Ergebnissen bestätigen. / Hypertrophic Cardiomyopathy is disease of the cardiac muscle which results in an asymmetric hypertrophy especially of the interventricularseptum of the heart. It is transmitted in an autosomal dominant way. The symptoms are unspecific reaching from dyspnoe to syncopes. Sometimes the sudden death is the first manifestation of the disease. Molekular genetic researches showed that in the patients genes Mutations in proteins of the sarkomer were detectable. Two of them are alpha-Tropomyosin and beta-Myosin Heavy Chain. We examined 45 unrelated Patient of the existence of Mutations in alpha-Tropomyosin and beta-Myosin Heavy Chain. We used following Examinations: PCR, SSCP, Sequencing. A mutation in the beta-Myosin Heavy Chain were found in 6 Patients (13%), non in alpha-Tropomyosin. Generally mutations are expected in 35% in beta-Myosin Heavy Chain and 3% in alpha-Tropomyosin. Our patients seem to have mutations in genes we did not examine in this study. We detected Mutations in: Exon 13 Arg403Trp and Val411Ile, Exon 19 Arg719Trp, Exon 20 Ile736Thr, Exon 21 Leu796Phe und Exon 23 Cys905Phe. Mutation Arg 403Trp and Arg719Trp have been known in this form before, the others were new. As the phenotypes of our patients were heterogenous and not significantly to be distinguished we looked for risk factors for sudden death as described by Burn et al. 1997 within our group of patients with mutations. Five Persons showed risk factors as discribed: Exon 13 Arg403Trp and Val411Ile, Exon 19 Arg719Trp, Exon 20 Ile736Thr, Exon 21 Leu796Phe. The person with the mutation Exon 23 Cys905Phe showed no risk factors for sudden death. Our results correlate with those of earlier studies. The patient with the mutation Exon 23 Cys905Phe was classified as a low risk patient while the other mutations correlate with a further high risk.

No description available.

Tropomyosins(Tm) are a group of proteins that regulate the actin filaments in both muscle and non-muscle cells. In mammalian cells four Tm species are found: α-Tm (fast) encoded by α-Tm /TPM1 gene, β-Tm, encoded by β-Tm/ TPM2 gene, α-Tm (slow) encoded by γTm gene/ TPM3 and δ-Tm encoded by δTm / TPM4gene. Mutations in Tm are linked to many cardiac and skeletal diseases like hypertrophic cardiac myopathy (TPM1 and TPM2), familial cardiac myopathy (TPM1) and skeletal diseases like nemaline myopathy (TPM2 and TPM3) along with other sarcomere proteins. The hypothesis on which this study is based is, the isoform composition in both muscle and non-muscle cells adapts in response to disease and physiological changes. A significant part of that adaptation is changes in the thin filament protein isoforms expressed and the post translational modifications of these proteins. In this study Tpm3.12st isoform of γTm and other striated muscle tropomyosin isoforms (Tpm1 and Tpm2) and a non-muscle Tmp4 were characterised using a variety of techniques. The aim was to enhance our understanding of the role of tropomyosin interactions in regards to its efficiency of actin binding capacity as well as its effect on actin polymerisation. Human tropomyosin 3 (Tpm3.12st) was expressed in E. coli to produce recombinant protein with three N-terminal sequence variants (Met, MM and (M)ASM). The proteins were characterised for their binding affinity with actin as this isoform has not been well characterised so far. Its properties are compared with other striated muscle tropomyosin Tpm1.1st and Tpm2.2st and non-muscle Tpm4.1cy. The proteins were purified through ion exchange chromatography and the purity was checked by using SDS-PAGE and UV spectrometry. The molecular weights of the recombinant proteins produced were confirmed by mass spectrometry. Cosedimentation assays were performed for their actin binding affinity using ultracentrifugation. The variant of Tpm3.12st with AS N-terminal extension was found to have similar actin affinity to Tpm1.1st in the range of 0.1-0.8 μM (half saturation). However the variants with Met and MM N-termini bound to actin weakly with high half saturation concentration of ~ 6 μM and ~8 μM tropomyosin respectively. Measurement of actin polymerisation kinetics showed it is affected in presence of tropomyosin. From this study it is shown that tropomyosin accelerates the initiation step in actin polymerisation with varying differences within the isoforms in contrast to several previous studies. There have been very few studies of the effect of tropomyosin on actin polymerisation in the last two decades. This work shows that tropomyosin isoforms have a large and variable role in controlling actin polymerisation and understanding tropomyosin function will need further investigation in this area. This study also developed an ELISA screening method using monoclonal antibodies for identification and quantification of Tpm3.12st which was tested against all the four tropomyosin isoforms. None of the twelve antibodies studied showed reactivity only with Tpm3.12st. From the data analysed it is deduced the amino acid residues in the region of 24-43 shows the prospect of designing a monoclonal antibody specific to Tpm3.12st isoform. Accurate quantification of tropomyosin isoforms is key to understanding their function and the effects of modulation of isoform composition in health and disease. A reverse phase liquid chromatography method was developed which is compatible with the analysis of the thin filament proteome using top-down mass spectrometry. Reverse phase liquid chromatography (RPLC) is one of the most popular methods used in mass spectrometry analysis where proteins are separated based on their hydrophobicity. The RPLC method developed in this study gives an efficient separation of major thin filament proteins along with small soluble proteins that is compatible to use for top down mass spectrometry for identification and quantification of proteins, PTMs and isoform composition. With a minimum amount of 2 mg of tissue using chicken and mouse heart and skeletal muscle samples a buffer system was optimized to extract thin filament proteins. With the optimized RPLC method actin, tropomyosin and troponin complex subunits (TnC, TnI and TnI) were successfully separated and the proteins were identified using SDS-page by comparison with the previous research results. This novel method of extraction and the optimised RPLC method will provide a “bird’s eye view” of thin filament proteome providing information of PTMs of all the proteins together within one single extraction, reducing the time for analysis and the sample size. This has the potential to give insight into tissue, muscle and heart adaptations that could act as a prognostic indicator.

Objetivo: O objetivo desse estudo foi avaliar alterações na resposta clínica e imunológica ao camarão após a imunoterapia com Dermatophagoides pteronyssinus. Métodos: Selecionou-se 35 indivíduos alérgicos a Dermatophagoides pteronyssinus (Der p), os quais foram submetidos a testes cutâneos de leitura imediata para ácaros, baratas, camarão, tropomiosina recombinante, além de cão, gato e fungos. A detecção de IgE espcífica in vitro foi feita para o ácaro, camarão, barata americana e para suas tropomiosinas. Em todos, avaliou-se reatividade clínica ao camarão através de provocação oral. Dez pacientes foram alocados para o grupo controle, e 25 foram submetidos à imunoterapia alérgeno específica para o ácaro. Os testes cutâneos e a dosagem de IgE sérica específica foram repetidas após a indução da imunoterapia, e após 1 ano do início. A reatividade clínica ao camarão foi reavaliada no final do estudo pela provocação oral. Resultados: No grupo dos pacientes que foram submetidos à imunoterapia, observamos diminuição na reatividade nos testes cutâneos e dosagem de IgE específica para Der p, camarão e tropomiosina recombinante. Dos 10 pacientes com testes cutâneos positivos para camarão, 4 foram negativos na dosagem após um ano de imunoterapia (p= 0,04). Quanto à dosagem sérica de IgE para camarão, dos 9 positivos no início, 6 ficaram negativos (p= 0,014). Nenhum paciente submetido a imunoterapia desenvolveu nova sensibilização para camarão. Não houve alteração na reatividade clínica ao camarão após imunoterapia. Conclusão: A imunoterapia para Dermatophagoides pteronyssinus foi acompanhada de diminuição da reatividade imunológica para camarão e clinicamente não houve alteração da sensibilidade a camarão / Objective: The objective of this study was to determine changes in clinical and immunological response to shrimp after immunotherapy with Dermatophagoides pteronyssinus. Methods: We studied 35 allergic subjects to Dermatophagoides pteronyssinus (Der p), submitted to skin tests to mites, cockroach, shrimp, recombinant tropomyosin, and dog, cat and fungi. The detection of serum specific IgE was performed to mite, shrimp, and tropomyosin from American cockroach. In all patients, the clinical reactivity to shrimp was assessed through oral challenge. Ten patients were allocated to the control group, and 25 were submitted to immunotherapy for mite. Skin tests and determination of serum specific IgE were repeated after the induction of immunotherapy (3-4 months) and 1 year after of beginning of the treatment. The clinical reactivity to shrimp was assessed again at the end of the study by oral challenge. Results: In the group of patients who were undergoing immunotherapy, we observed decreased reactivity in the skin tests and specific IgE levels to Der p, shrimp and recombinant tropomyosin. Among the 10 patients with positive skin tests to shrimp, 4 were negative when assessed after one year of immunotherapy (p = 0.04). About serum specific IgE to shrimp, from the 9 positive reactors in the beginning of treatment, 6 became negative (p= 0.014). There was no change in clinical reactivity to shrimp after immunotherapy. Conclusion: The immunotherapy for Dermatophagoides pteronyssinus was accompanied by decreased immune reactivity to shrimp and clinically there was no change in sensitivity to shrimp

Objetivo: O objetivo desse estudo foi avaliar alterações na resposta clínica e imunológica ao camarão após a imunoterapia com Dermatophagoides pteronyssinus. Métodos: Selecionou-se 35 indivíduos alérgicos a Dermatophagoides pteronyssinus (Der p), os quais foram submetidos a testes cutâneos de leitura imediata para ácaros, baratas, camarão, tropomiosina recombinante, além de cão, gato e fungos. A detecção de IgE espcífica in vitro foi feita para o ácaro, camarão, barata americana e para suas tropomiosinas. Em todos, avaliou-se reatividade clínica ao camarão através de provocação oral. Dez pacientes foram alocados para o grupo controle, e 25 foram submetidos à imunoterapia alérgeno específica para o ácaro. Os testes cutâneos e a dosagem de IgE sérica específica foram repetidas após a indução da imunoterapia, e após 1 ano do início. A reatividade clínica ao camarão foi reavaliada no final do estudo pela provocação oral. Resultados: No grupo dos pacientes que foram submetidos à imunoterapia, observamos diminuição na reatividade nos testes cutâneos e dosagem de IgE específica para Der p, camarão e tropomiosina recombinante. Dos 10 pacientes com testes cutâneos positivos para camarão, 4 foram negativos na dosagem após um ano de imunoterapia (p= 0,04). Quanto à dosagem sérica de IgE para camarão, dos 9 positivos no início, 6 ficaram negativos (p= 0,014). Nenhum paciente submetido a imunoterapia desenvolveu nova sensibilização para camarão. Não houve alteração na reatividade clínica ao camarão após imunoterapia. Conclusão: A imunoterapia para Dermatophagoides pteronyssinus foi acompanhada de diminuição da reatividade imunológica para camarão e clinicamente não houve alteração da sensibilidade a camarão / Objective: The objective of this study was to determine changes in clinical and immunological response to shrimp after immunotherapy with Dermatophagoides pteronyssinus. Methods: We studied 35 allergic subjects to Dermatophagoides pteronyssinus (Der p), submitted to skin tests to mites, cockroach, shrimp, recombinant tropomyosin, and dog, cat and fungi. The detection of serum specific IgE was performed to mite, shrimp, and tropomyosin from American cockroach. In all patients, the clinical reactivity to shrimp was assessed through oral challenge. Ten patients were allocated to the control group, and 25 were submitted to immunotherapy for mite. Skin tests and determination of serum specific IgE were repeated after the induction of immunotherapy (3-4 months) and 1 year after of beginning of the treatment. The clinical reactivity to shrimp was assessed again at the end of the study by oral challenge. Results: In the group of patients who were undergoing immunotherapy, we observed decreased reactivity in the skin tests and specific IgE levels to Der p, shrimp and recombinant tropomyosin. Among the 10 patients with positive skin tests to shrimp, 4 were negative when assessed after one year of immunotherapy (p = 0.04). About serum specific IgE to shrimp, from the 9 positive reactors in the beginning of treatment, 6 became negative (p= 0.014). There was no change in clinical reactivity to shrimp after immunotherapy. Conclusion: The immunotherapy for Dermatophagoides pteronyssinus was accompanied by decreased immune reactivity to shrimp and clinically there was no change in sensitivity to shrimp