Characterization and location of Src-dependent tyrosine phosphorylation in rat brain mitochondria

Analysis of protein phosphorylation in highly purified rat brain mitochondria revealed the presence of several alkali-stable phosphoproteins whose phosphorylation markedly increases upon treatment with peroxovanadate and Mn(2+), a property indicating tyrosine phosphorylation. These include three prominent bands, with apparent sizes of 50, 60, and 75 kDa, which are detectable by anti-phosphotyrosine. Tyrosine phosphorylation disappears when mitochondria are treated with PP2, an inhibitor of the Src kinase family, suggesting the presence of members of this family in rat brain mitochondria. Immunoblotting and immunoprecipitation assays of mitochondrial lysates confirmed the presence of Fyn, Src and Lyn kinases, as well as Csk, a protein kinase which negatively controls the activity of the Src kinase family. Results show that tyrosine-phosphorylated proteins are membrane-bound and that they are located on the inner surface of the outer membrane and/or the external surface of the inner membrane. Instead, Src tyrosine kinases are mainly located in the intermembrane space - in particular, as revealed by immunogold experiments for Lyn kinase, in the cristal lumen. Rat brain mitochondria were also found to possess a marked level of tyrosine phosphatase activity, strongly inhibited by peroxovanadate.     

Effect of peroxides on spermine transport in rat brain and liver mitochondria

The polyamine spermine is transported into the matrix of various types of mitochondria by a specific uniporter system identified as a protein channel. This mechanism is regulated by the membrane potential; other regulatory effectors are unknown. This study analyzes the transport of spermine in the presence of peroxides in both isolated rat liver and brain mitochondria, in order to evaluate the involvement of the redox state in this mechanism, and to compare its effect in both types of mitochondria. In liver mitochondria peroxides are able to inhibit spermine transport. This effect is indicative of redox regulation by the transporter, probably due to the presence of critical thiol groups along the transport pathway, or in close association with it, with different accessibility for the peroxides and performing different functions. In brain mitochondria, peroxides have several effects, supporting the hypothesis of a different regulation of spermine transport. The fact that peroxovanadate can inhibit tyrosine phosphatases in brain mitochondria suggests that mitochondrial spermine transport is regulated by tyrosine phosphorylation in this organ. In this regard, the evaluation of spermine transport in the presence of Src inhibitors suggests the involvement of Src family kinases in this process. It is possible that phosphorylation sites for Src kinases are present in the channel pathway and have an inhibitory effect on spermine transport under regulation by Src kinases. The results of this study suggest that the activity of the spermine transporter probably depends on the redox and/or tyrosine phosphorylation state of mitochondria, and that its regulation may be different in distinct organs.     

Src family protein tyrosine kinases induce autoactivation of Bruton's tyrosine kinase.

Bruton's tyrosine kinase (Btk) is tyrosine phosphorylated and enzymatically activated following ligation of the B-cell antigen receptor. These events are temporally regulated, and Btk activation follows that of various members of the Src family of protein tyrosine kinases, thus raising the possibility that Src kinases participate in the Btk activation process. We have evaluated the mechanism underlying Btk enzyme activation and have explored the potential regulatory relationship between Btk and Src protein kinases. We demonstrate in COS transient-expression assays that Btk can be activated through intramolecular autophosphorylation at tyrosine 551 and that Btk autophosphorylation is required for Btk catalytic functions. Coexpression of Btk with members of the Src family of protein tyrosine kinases, but not Syk, led to Btk tyrosine phosphorylation and activation. Using a series of point mutations in Blk (a representative Src protein kinase) and Btk, we show that Src kinases activate Btk through an indirect mechanism that requires membrane association of the Src enzymes as well as functional Btk SH3 and SH2 domains. Our results are compatible with the idea that Src protein tyrosine kinases contribute to Btk activation by indirectly stimulating Btk intramolecular autophosphorylation.     

Cloning of the doding cDNA sequence of the gene for Csk tyrosine kinase from human lymphocytes

Tyrosine kinases of the Csk family play an important role in cell growth regulation and normal cell differentiation. They are also involved in carcinogenesis as oncoproteins. The main function of these tyrosine kinases is phosphorylation of tyrosine kinases of the Src family at their C-terminal regions to negatively regulate their activity. Disturbance of csk expression increases the Src tyrosine kinase activity. The full-length coding sequence of the csk cDNA was cloned from human lymphocytes. The 1624-bp cDNA consists of 12 exons and encodes a protein that has conserved SH2 and SH3 domains and is similar to human Csk tyrosine kinase by 99%. The full-length cDNA can be used to analyze the csk structure in normal or illdefined human cells.     

Identification of the flavoprotein of succinate dehydrogenase and aconitase as in vitro mitochondrial substrates of Fgr tyrosine kinase

Overlooked until recently, mitochondrial protein phosphorylation is now emerging as a key post-translational mechanism in the regulation of mitochondrial functions. In particular, tyrosine phosphorylation represents a promising field to discover new mechanisms of bioenergetic regulation. Tyrosine kinases belonging to the Src kinase family have been observed in mitochondrial compartments, however their substrates are almost unknown. Here, we provide evidence that the flavoprotein of succinate dehydrogenase and aconitase are "in vitro" substrates of Fgr tyrosine kinase. Fgr phosphorylates flavoprotein of succinate dehydrogenase at Y535 and Y596 and aconitase at Y71, Y544 and Y665. The significance of these findings is discussed.     

Tyrosine phosphorylation of the N-methyl-D-aspartate receptor by exogenous and postsynaptic density-associated Src-family kinases

Phosphorylation of the NMDA receptor by Src-family tyrosine kinases has been implicated in the regulation of receptor function. We have investigated the tyrosine phosphorylation of NMDA receptor subunits NR2A and NR2B by exogenous Src and Fyn and compared this to phosphorylation by tyrosine kinases associated with the postsynaptic density (PSD). Phosphorylation of the receptor by exogenous Src and Fyn was dependent upon initial binding of the kinases to PSDs via their SH2-domains. Src and Fyn phosphorylated similar sites in NR2A and NR2B, tryptic peptide mapping identifying seven and five major tyrosine-phosphorylated peptides derived from NR2A and NR2B, respectively. All five tyrosine phosphorylation sites on NR2B were localized to the C-terminal, cytoplasmic domain. Phosphorylation of NR2B by endogenous PSD tyrosine kinases yielded only three tyrosine-phosphorylated tryptic peptides, two of which corresponded to Src phosphorylation sites, and one of which was novel. Phosphorylation-site specific antibodies identified NR2B Tyr1472 as a phosphorylation site for intrinsic PSD tyrosine kinases. Phosphorylation of this site was inhibited by the Src-family-specific inhibitor PP2. The results identify several potential phosphorylation sites for Src in the NMDA receptor, and indicate that not all of these sites are available for phosphorylation by kinases located within the structural framework of the PSD.     

Src tyrosine kinase-dependent migratory effects of antithrombin in leukocytes

Tyrosine kinases are known to play a critical role in the regulation of leukocyte function. Antithrombin mediates its effects via syndecan-4 which is known to be linked to the Src tyrosine kinases. In this study, we investigated the role of Src tyrosine kinases in antithrombin-regulated leukocyte migration and Src tyrosine kinase phosphorylation in response to stimulation with antithrombin. Neutrophils and monocytes obtained from forearm venous blood were pre-treated by various Src-family selective tyrosine kinase inhibitors with or without antithrombin followed by washing and assessment of their migratory response toward antithrombin, interleukin-8, or RANTES using Boyden microchemotaxis chambers. Activation status of the two major Src tyrosine kinase phosphorylation sides Tyr416 and Tyr527 was tested using Western blot analysis. Dose-dependent reversal of the antithrombin-mediated effects on neutrophil and monocyte migration was induced by the selective Src kinase inhibitors PP1 and PP2. In Western blot analyses, antithrombin increased Tyr416 and decreased Tyr527 phosphorylation of Src tyrosine kinases in a time- and dose-dependent manner. Moreover, co-incubation with antithrombin lowered the level of RANTES-induced Tyr416 phosphorylation. Therefore, Src tyrosine kinases linked to signaling of antithrombin-binding sites on leukocytes may play an important role in modulating effects on cells function.     

Phosphorylation of a Src kinase at the autophosphorylation site in the absence of Src kinase activity

Exposure of cells to oxidants increases the phosphorylation of the Src family tyrosine protein kinase Lck at Tyr-394, a conserved residue in the activation loop of the catalytic domain. Kinase-deficient Lck expressed in fibroblasts that do not express any endogenous Lck has been shown to be phosphorylated at Tyr-394 following H(2)O(2) treatment to an extent indistinguishable from that seen with wild type Lck. This finding indicates that a kinase other than Lck itself is capable of phosphorylating Tyr-394. Because fibroblasts express other Src family members, it remained to be determined whether the phosphorylation of Tyr-394 was carried out by another Src family kinase or by an unrelated tyrosine protein kinase. We examined here whether Tyr-394 in kinase-deficient Lck was phosphorylated following exposure of cells devoid of endogenous Src family kinase activity to H(2)O(2). Strikingly, treatment of such cells with H(2)O(2) led to the phosphorylation of Tyr-394 to an extent identical to that seen with wild type Lck, demonstrating that Src family kinases are not required for H(2)O(2)-induced phosphorylation of Lck. Furthermore, this efficient phosphorylation of Lck at Tyr-394 in non-lymphoid cells suggests the existence of an ubiquitous activator of Src family kinases.     

Src和Abl酪氨酸蛋白激酶家族参与病原微生物感染的研究进展

Src和Abl家族激酶属于非受体型酪氨酸激酶(Nonreceptor tyrosine kinase,NRTK)家族重要成员,广泛存在于各种细胞中,参与细胞内信号传递并调节细胞生理过程,它们在维持细胞、组织和器官稳态功能中发挥着至关重要的作用。研究表明,Src和Abl家族激酶通过多种机制参与病原微生物的感染(如与病原微生物的脯氨酸基序-PXXP互作)。因此,从Src和Abl家族激酶角度出发探究病原微生物感染机制逐渐成为一个热点。本文就Src和Abl家族激酶的结构特点以及参与病原微生物感染的研究报道进行综述,以期为病原微生物感染的致病机制、防控和药物研发提供参考。     

Complex Formation with Focal Adhesion Kinase: A Mechanism to Regulate Activity and Subcellular Localization of Src Kinases

Tyrosine phosphorylation of focal adhesion kinase (FAK) creates a high-affinity binding site for the src homology 2 domain of the Src family of tyrosine kinases. Assembly of a complex between FAK and Src kinases may serve to regulate the subcellular localization and the enzymatic activity of members of the Src family of kinases. We show that simultaneous overexpression of FAK and pp60(c-src) or p59(fyn) results in the enhancement of the tyrosine phosphorylation of a limited number of cellular substrates, including paxillin. Under these conditions, tyrosine phosphorylation of paxillin is largely cell adhesion dependent. FAK mutants defective for Src binding or focal adhesion targeting fail to cooperate with pp60(c-src) or p59(fyn) to induce paxillin phosphorylation, whereas catalytically defective FAK mutants can direct paxillin phosphorylation. The negative regulatory site of pp60(c-src) is hypophosphorylated when in complex with FAK, and coexpression with FAK leads to a redistribution of pp60(c-src) from a diffuse cellular location to focal adhesions. A FAK mutant defective for Src binding does not effectively induce the translocation of pp60(c-src) to focal adhesions. These results suggest that association with FAK can alter the localization of Src kinases and that FAK functions to direct phosphorylation of cellular substrates by recruitment of Src kinases.     

alpha-synuclein is phosphorylated by members of the Src family of protein-tyrosine kinases

alpha-Synuclein (alpha-Syn) is implicated in the pathogenesis of Parkinson's Disease, genetically through missense mutations linked to early onset disease and pathologically through its presence in Lewy bodies. alpha-Syn is phosphorylated on serine residues; however, tyrosine phosphorylation of alpha-Syn has not been established (, ). A comparison of the protein sequence between Synuclein family members revealed that all four tyrosine residues of alpha-Syn are conserved in all orthologs and beta-Syn paralogs described to date, suggesting that these residues may be of functional importance (). For this reason, experiments were performed to determine whether alpha-Syn could be phosphorylated on tyrosine residue(s) in human cells. Indeed, alpha-Syn is phosphorylated within 2 min of pervanadate treatment in alpha-Syn-transfected cells. Tyrosine phosphorylation occurs primarily on tyrosine 125 and was inhibited by PP2, a selective inhibitor of Src protein-tyrosine kinase (PTK) family members at concentrations consistent with inhibition of Src function (). Finally, we demonstrate that alpha-Syn can be phosphorylated directly both in cotransfection experiments using c-Src and Fyn expression vectors and in in vitro kinase assays with purified kinases. These data suggest that alpha-Syn can be a target for phosphorylation by the Src family of PTKs.     

Abl collaborates with Src family kinases to stimulate actin-based motility of vaccinia virus

Local activation of Src at the plasma membrane by extracellular vaccinia virus results in a signalling cascade that acts to stimulate actin polymerization beneath the virus to enhance its cell-to-cell spread. Initiation of this signalling cascade involves Src-mediated phosphorylation of tyrosine 112 and 132 of the viral membrane protein A36R. Here we show that recruitment of Src is dependent on its myristoylation and an interaction with A36R upstream of tyrosine 112 and 132. We further show that Src, Fyn and Yes have unique specificities towards these tyrosine residues. Using cell lines deficient in Src, Fyn and Yes, we demonstrate that multiple Src family members can stimulate vaccinia-induced actin polymerization and also uncover a role for Abl family kinases. Additionally, Abl and Arg are able to phosphorylate A36R in vitro and are recruited to vaccinia-induced actin tails. The ability of multiple families of tyrosine kinases to directly phosphorylate A36R ensures robust cell-to-cell spread of vaccinia virus will occur under a variety of cellular conditions.     

Identification of Tyr900 in the kinase domain of c-Kit as a Src-dependent phosphorylation site mediating interaction with c-Crk

We have previously demonstrated that ligand-stimulation of c-Kit induces phosphorylation of Tyr568 and Tyr570 in the juxtamembrane region of the receptor, leading to recruitment, phosphorylation and activation of members of the Src family of tyrosine kinases. In this paper, we demonstrate that members of the Src family of tyrosine kinases are able to phosphorylate c-Kit selectively on one particular tyrosine residue, Tyr900, located in the second part of the tyrosine kinase domain. In order to identify potential docking partners of Tyr900, a synthetic phosphopeptide corresponding to the amino acid sequence surrounding Tyr900 was used as an affinity matrix. By use of MALDI-TOF mass spectrometry, CrkII was identified as a protein that specifically bound to Tyr900 in a phosphorylation dependent manner, possibly via the p85 subunit of PI3-kinase. Expression of a mutant receptor where Tyr900 had been replaced with a phenylalanine residue (Y900F) resulted in a receptor with reduced ability to phosphorylate CrkII. Together these data support a model where c-Src phosphorylates the receptor, thereby creating docking sites for SH2 domain containing proteins, leading to recruitment of Crk to the receptor.     

Regulation of a transient receptor potential (TRP) channel by tyrosine phosphorylation. SRC family kinase-dependent tyrosine phosphorylation of TRPV4 on TYR-253 mediates its response to hypotonic stress

The recently identified transient receptor potential (TRP) channel family member, TRPV4 (formerly known as OTRPC4, VR-OAC, TRP12, and VRL-2) is activated by hypotonicity. It is highly expressed in the kidney as well as blood-brain barrier-deficient hypothalamic nuclei responsible for systemic osmosensing. Apart from its gating by hypotonicity, little is known about TRPV4 regulation. We observed that hypotonic stress resulted in rapid tyrosine phosphorylation of TRPV4 in a heterologous expression model and in native murine distal convoluted tubule cells in culture. This tyrosine phosphorylation was sensitive to the inhibitor of Src family tyrosine kinases, PP1, in a dose-dependent fashion. TRPV4 associated with Src family kinases by co-immunoprecipitation studies and confocal immunofluorescence microscopy, and this interaction required an intact Src family kinase SH2 domain. One of these kinases, Lyn, was activated by hypotonic stress and phosphorylated TRPV4 in an immune complex kinase assay and an in vitro kinase assay using recombinant Lyn and TRPV4. Transfection of wild-type Lyn dramatically potentiated hypotonicity-dependent TRPV4 tyrosine phosphorylation whereas dominant negative-acting Lyn modestly inhibited it. Through mutagenesis studies, the site of tonicity-dependent tyrosine phosphorylation was mapped to Tyr-253, which is conserved across all species from which TRPV4 has been cloned. Importantly, point mutation of Tyr-253 abolished hypotonicity-dependent channel activity. In aggregate, these data indicate that hypotonic stress results in Src family tyrosine kinase-dependent tyrosine phosphorylation of the tonicity sensor TRPV4 at residue Tyr-253 and that this residue is essential for channel function in this context. This is the first example of direct regulation of TRP channel function through tyrosine phosphorylation.     

Prothoracicotropic hormone induces tyrosine phosphorylation in prothoracic glands of the silkworm,Bombyx mori

In the present study, we investigated the tyrosine phosphorylation of Bombyx mori prothoracic glands using phosphotyrosine‐specific antibodies and Western blot analysis. Results showed that prothoracicotropic hormone (PTTH) stimulates a rapid increase in tyrosine phosphorylation of at least 2 proteins in prothoracic glands, one of which was identified as extracellular signal‐regulated kinase (ERK). The phosphorylation of another 120‐kDa protein showed dose‐ and time‐dependent stimulation by PTTH in vitro. In vitro activation of tyrosine phosphorylation was also verified by in vivo experiments: injection of PTTH into day‐6 last‐instar larvae greatly increased tyrosine phosphorylation. Treatment of prothoracic glands with the protein tyrosine phosphatase inhibitor, sodium orthovanadate, also resulted in tyrosine phosphorylation of several proteins and increased ecdysteroidogenesis. The PTTH‐stimulated phosphorylation of the 120‐kDa protein was markedly attenuated by genistein, a broad‐spectrum tyrosine kinase inhibitor, but not by HNMPA‐(AM)₃, a specific inhibitor of insulin receptor tyrosine kinase. PP2, a more‐selective inhibitor of the Src‐family tyrosine kinases, partially inhibited PTTH‐stimulated tyrosine phosphorylation, but not ecdysteroidogenesis. This result implies the possibility that in addition to ERK, the phosphorylation of the 120‐kDa protein, which is not Src‐family tyrosine kinase, is likely also involved in PTTH‐stimulated ecdysteroidogenesis in B. mori. © 2010 Wiley Periodicals, Inc.     

Src redox regulation: there is more than meets the eye

Src-family kinases are critically involved in the control of cytoskeleton organization and in the generation of integrin-dependent signaling responses, inducing tyrosine phosphorylation of many signaling and cytoskeletal proteins. Activity of the Src family of tyrosine kinases is tightly controlled by inhibitory phosphorylation of a carboxy-terminal tyrosine residue, inducing an inactive conformation through binding with its SH2 domain. Dephosphorylation of C-ter tyrosine, as well as its deletion of substitution with phenylalanine in oncogenic Src kinases, leads to autophosphorylation at a tyrosine in the activation loop, thereby leading to enhanced Src activity. Beside this phophorylation/dephosphorylation circuitry, cysteine oxidation has been recently reported as a further mechanism of enzyme activation. Mounting evidence describes Src activation via its redox regulation as a key outcome in several circumstances, including growth factor and cytokines signaling, integrin-mediated cell adhesion and motility, membrane receptor cross-talk as well in cell transformation and tumor progression. Among the plethora of data involving Src kinase in physiological and pathophysiological processes, this review will give emphasis to the redox component of the regulation of this master kinase.     

Syk-dependent phosphorylation of CLEC-2: a novel mechanism of hem-immunoreceptor tyrosine-based activation motif signaling

The C-type lectin-like receptor CLEC-2 signals via phosphorylation of a single cytoplasmic YXXL sequence known as a hem-immunoreceptor tyrosine-based activation motif (hemITAM). In this study, we show that phosphorylation of CLEC-2 by the snake toxin rhodocytin is abolished in the absence of the tyrosine kinase Syk but is not altered in the absence of the major platelet Src family kinases, Fyn, Lyn, and Src, or the tyrosine phosphatase CD148, which regulates the basal activity of Src family kinases. Further, phosphorylation of CLEC-2 by rhodocytin is not altered in the presence of the Src family kinase inhibitor PP2, even though PLCγ2 phosphorylation and platelet activation are abolished. A similar dependence of phosphorylation of CLEC-2 on Syk is also seen in response to stimulation by an IgG mAb to CLEC-2, although interestingly CLEC-2 phosphorylation is also reduced in the absence of Lyn. These results provide the first definitive evidence that Syk mediates phosphorylation of the CLEC-2 hemITAM receptor with Src family kinases playing a critical role further downstream through the regulation of Syk and other effector proteins, providing a new paradigm in signaling by YXXL-containing receptors.     

Regulation of Ly49D/DAP12 signal transduction by Src-family kinases and CD45

Activating, DAP12-coupled members of the Ly-49 family of NK cell receptors help control viral infections in mice. However, the kinases and/or phosphatases mediating tyrosine phosphorylation of Ly-49D-associated DAP12 have not been elucidated. In this study, we show for the first time that Src family tyrosine kinases are physically and functionally associated with Ly-49D/DAP12 signaling in murine NK cells. Specifically, we demonstrate the following: 1) inhibition of Src family kinases suppresses DAP12 phosphorylation and downstream DAP12 signals; 2) both Fyn and Lck are capable of phosphorylating DAP12; and 3) both kinases coimmunoprecipitate with the Ly-49D/DAP12 complex in NK cells. Although we detect enhanced phosphorylation of Fyn upon Ly-49D cross-linking in NK cells, Ly-49D-mediated events in both Fyn-/- and Fyn/Lck-/- mice appear normal, reinforcing the theme of redundancy in the ability of Src family kinases to initiate activation events. In contrast to disruption of specific Src family enzymes, Ly-49D/DAP12-mediated calcium mobilization and cytokine production by CD45 null NK cells are defective. Although others have ascribed the effects of CD45 mutation solely on the suppression of Src family activity, we demonstrate in this study that DAP12 is hyperphosphorylated in CD45 null NK cells, resulting in uncoordinated tyrosine-mediated signaling upon Ly-49D ligation. Therefore, although our data are consistent with a Src kinase activity proximally within DAP12 signaling, DAP12 also appears to be a substrate of CD45, suggesting a more complex role for this phosphatase than has been reported previously.     

Introduction of p130cas signaling complex formation upon integrin-mediated cell adhesion: a role for Src family kinases.

Integrin-mediated cell adhesion triggers intracellular signaling cascades, including tyrosine phosphorylation of intracellular proteins. Among these are the focal adhesion proteins p130cas (Cas) and focal adhesion kinase (FAK). Here we identify the kinase(s) mediating integrin-induced Cas phosphorylation and characterize protein-protein interactions mediated by phosphorylated Cas. We found that expression of a constitutively active FAK in fibroblasts results in a consecutive tyrosine phosphorylation of Cas. This effect required the autophosphorylation site of FAK, which is a binding site for Src family kinases. Integrin-mediated phosphorylation of Cas was not, however, compromised in fibroblasts lacking FAK. In contrast, adhesion-induced tyrosine phosphorylation of Cas was reduced in cells lacking Src, whereas enhanced phosphorylation of Cas was observed Csk- cells, in which Src kinases are activated. These results suggest that Src kinases are responsible for the integrin-mediated tyrosine phosphorylation of Cas. FAK seems not to be necessary for phosphorylation of Cas, but when autophosphorylated, FAK may recruit Src family kinases to phosphorylate Cas. Cas was found to form complexes with Src homology 2 (SH2) domain-containing signaling molecules, such as the SH2/SH3 adapter protein Crk, following integrin-induced tyrosine phosphorylation. Guanine nucleotide exchange factors C3G and Sos were found in the Cas-Crk complex upon integrin ligand binding. These observations suggest that Cas serves as a docking protein and may transduce signals to downstream signaling pathways following integrin-mediated cell adhesion.     

Interaction between the SH3 domain of Src family kinases and the proline-rich motif of HTLV-1 p13: a novel mechanism underlying delivery of Src family kinases to mitochondria

The association of the SH3 (Src homology 3) domain of SFKs (Src family kinases) with protein partners bearing proline-rich motifs has been implicated in the regulation of SFK activity, and has been described as a possible mechanism of relocalization of SFKs to subcellular compartments. We demonstrate in the present study for the first time that p13, an accessory protein encoded by the HTLV-1 (human T-cell leukaemia virus type?1), binds the SH3 domain of SFKs via its C-terminal proline-rich motif, forming a stable heterodimer that translocates to mitochondria by virtue of its N-terminal mitochondrial localization signal. As a result, the activity of SFKs is dramatically enhanced, with a subsequent increase in mitochondrial tyrosine phosphorylation, and the recognized ability of p13 to insert itself into the inner mitochondrial membrane and to perturb the mitochondrial membrane potential is abolished. Overall, the present study, in addition to confirming that the catalytic activity of SFKs is modulated by interactors of their SH3 domain, leads us to hypothesize a general mechanism by which proteins bearing a proline-rich motif and a mitochondrial localization signal at the same time may act as carriers of SFKs into mitochondria, thus contributing to the regulation of mitochondrial functions under various pathophysiological conditions.