The functional cooperation of MAP1A heavy chain and light chain 2 in the binding of microtubules

Microtubule-associated protein 1A (MAP1A) is a high-molecular-weight protein that is comprised of a heavy chain and a light chain (LC2) and is widely distributed along the microtubules in both mature neurons and glial cells. To illustrate the interaction among the MAP1A heavy chain, light chain, and microtubule, we prepared DNA constructs with Myc-, EGFP-, or DsRed-tags for full-length MAP1A DNA expressing whole MAP1A protein, two domains of MAP1A heavy chain, and light chain. Distribution patterns of various MAP1A domains as well as their interactions with microtubules were monitored in a non-neuronal COS7 and a neuronal Neuro2A cells. Our data revealed that a complete MAP1A protein, which contains both heavy chain and LC2, could be colocalized with microtubule networks not only in Neuro2A cells but also in transfected COS7 cells. Filamentous structures failed to be visualized along microtubules in COS7 cells transfected with MAP1A heavy chain or LC2 alone. Whereas, after introducing MAP1A heavy chain with LC2 into COS7 cells, both heavy chain and LC2 could be colocalized with microtubules. From our functional analysis, both MAP1A and its LC2 could protect microtubules against the challenge of nacodazol. Data collected from yeast two-hybrid assays of various MAP1A domains confirmed that the interaction of LC2 and NH2-terminal of MAP1A heavy chain is important for microtubule binding. From our analysis of MAP1A functional domains, we suggest that interactions between MAP1A heavy chain and LC2 are critical for the binding of microtubules.     

MAP1A light chain-2 interacts with GTP-RhoB to control epidermal growth factor (EGF)-dependent EGF receptor signaling

Rho GTPases have been implicated in the control of several cellular functions, including regulation of the actin cytoskeleton, cell proliferation, and oncogenesis. Unlike RhoA and RhoC, RhoB localizes in part to endosomes and controls endocytic trafficking. Using a yeast two-hybrid screen and a glutathione S-transferase pulldown assay, we identified LC2, the light chain of the microtubule-associated protein MAP1A, as a novel binding partner for RhoB. GTP binding and the 18-amino acid C-terminal hypervariable domain of RhoB are critical for its binding to MAP1A/LC2. Coimmunoprecipitation and immunofluorescence experiments showed that this interaction occurs in U87 cells. Down-regulation of MAP1A/LC2 expression decreased epidermal growth factor (EGF) receptor expression and modified the signaling response to EGF treatment. We concluded that MAP1A/LC2 is critical for RhoB function in EGF-induced EGF receptor regulation. Because MAP1A/LC2 is thought to function as an adaptor between microtubules and other molecules, we postulate that the RhoB and MAP1A/LC2 interactions facilitate endocytic vesicle trafficking and regulate the trafficking of signaling molecules.     

Microtubule-associated Proteins 1 (MAP1) Promote Human Immunodeficiency Virus Type I (HIV-1) Intracytoplasmic Routing to the Nucleus

After cell entry, HIV undergoes rapid transport toward the nucleus using microtubules and microfilaments. Neither the cellular cytoplasmic components nor the viral proteins that interact to mediate transport have yet been identified. Using a yeast two-hybrid screen, we identified four cytoskeletal components as putative interaction partners for HIV-1 p24 capsid protein: MAP1A, MAP1S, CKAP1, and WIRE. Depletion of MAP1A/MAP1S in indicator cell lines and primary human macrophages led to a profound reduction in HIV-1 infectivity as a result of impaired retrograde trafficking, demonstrated by a characteristic accumulation of capsids away from the nuclear membrane, and an overall defect in nuclear import. MAP1A/MAP1S did not impact microtubule network integrity or cell morphology but contributed to microtubule stabilization, which was shown previously to facilitate infection. In addition, we found that MAP1 proteins interact with HIV-1 cores both in vitro and in infected cells and that interaction involves MAP1 light chain LC2. Depletion of MAP1 proteins reduced the association of HIV-1 capsids with both dynamic and stable microtubules, suggesting that MAP1 proteins help tether incoming viral capsids to the microtubular network, thus promoting cytoplasmic trafficking. This work shows for the first time that following entry into target cells, HIV-1 interacts with the cytoskeleton via its p24 capsid protein. Moreover, our results support a role for MAP1 proteins in promoting efficient retrograde trafficking of HIV-1 by stimulating the formation of stable microtubules and mediating the association of HIV-1 cores with microtubules.     

Casein kinase 1 delta (CK1delta) interacts with the SNARE associated protein snapin

In this study we identified snapin as an interaction partner of the CK1 isoform delta (CK1delta) in the yeast two-hybrid system and localized the interacting domains of both proteins. The interaction of CK1delta with snapin was confirmed by co-immunoprecipitation. Snapin was phosphorylated by CK1delta in vitro. Both proteins localized in close proximity in the perinuclear region, wherein snapin was found to associate with membranes of the Golgi apparatus. The identification of snapin as a new substrate of CK1delta points towards a possible function for CK1delta in modulating snapin specific functions.     

Microtubule-associated protein 1S (MAP1S) bridges autophagic components with microtubules and mitochondria to affect autophagosomal biogenesis and degradation

The ubiquitously distributed MAP1S is a homologue of the exclusively neuronal distributed microtubule-associated protein 1A and 1B (MAP1A/B). They give rise to multiple isoforms through similar post-translational modification. Isoforms of MAP1S have been implicated in microtubule dynamics and mitotic abnormalities and mitotic cell death. Here we show that ablation of the Map1s gene in mice caused reduction in the B-cell CLL/lymphoma 2 or xL (Bcl-2/xL) and cyclin-dependent kinase inhibitor 1B (P27) protein levels, accumulation of defective mitochondria, and severe defects in response to nutritive stress, suggesting defects in autophagosomal biogenesis and clearance. Furthermore, MAP1S isoforms interacted with the autophagosome-associated light chain 3 of MAP1A/B (LC3), a homologue of yeast autophagy-related gene 8 (ATG8), and recruited it to stable microtubules in a MAP1S and LC3 isoform-dependent mode. In addition, MAP1S interacted with mitochondrion-associated leucine-rich PPR-motif containing protein (LRPPRC) that interacts with the mitophagy initiator and Parkinson disease-related protein Parkin. The three-way interactions of MAP1S isoforms with LC3 and microtubules as well as the interaction of MAP1S with LRPPRC suggest that MAP1S isoforms may play positive roles in integration of autophagic components with microtubules and mitochondria in both autophagosomal biogenesis and degradation. For the first time, our results clarify roles of MAP1S in bridging microtubules and mitochondria with autophagic and mitophagic initiation, maturation, trafficking, and lysosomal clearance. Defects in the MAP1S-regulated autophagy may impact heart disease, cancers, neurodegenerative diseases, and a wide range of other diseases.     

Human Microtubule-Associated Protein 1a (MAP1A) Gene: Genomic Organization, cDNA Sequence, and Developmental- and Tissue-Specific Expression

Microtubule-associated proteins (MAPs) regulate microtubule stability and play critical roles in neuronal development and the balance between neuronal plasticity and rigidity. MAP1a (HGMW-approved symbol MAP1A) stabilizes microtubules in postnatal axons. We describe human MAP1a's genomic organization and deduced cDNA and amino acid sequences. MAP1a is a single-copy gene spanning 10.5 kb. MAP1a coding sequence is contained in five exons. Translation begins in exon 3. Human MAP1a contains 2805 amino acids (predicted molecular weight 306.5 kDa) and is slightly larger than rat MAP1a (2774 amino acids). Like rat and bovine MAP1a, human MAP1a contains conserved tubulin binding motifs in the amino-terminal region. The carboxy-terminal portion contains a conserved pentadecapeptide that is present in the light chain portion of rat and bovine MAP1a/LC2 polyprotein. We show that human MAP1a gene expression occurs almost exclusively in the brain and that there is approximately 10-fold greater gene expression in adult brain compared to fetal brain. Strong, interspecies conservation between human and rat MAP1a cDNA and amino acid sequences indicates important relationships between MAP1a's function and its primary amino acid sequence.     

Microtubule Associated Protein 1b (MAP1B) Is a Marker of the Microtubular Cytoskeleton in Podocytes but Is Not Essential for the Function of the Kidney Filtration Barrier in Mice

Podocytes are essential for the function of the kidney glomerular filter. A highly differentiated cytoskeleton is requisite for their integrity. Although much knowledge has been gained on the organization of cortical actin networks in podocyte’s foot processes, less is known about the molecular organization of the microtubular cytoskeleton in primary processes and the cell body. To gain an insight into the organization of the microtubular cytoskeleton of the podocyte, we systematically analyzed the expression of microtubule associated proteins (Maps), a family of microtubules interacting proteins with known functions as regulator, scaffold and guidance proteins. We identified microtubule associated protein 1b (MAP1B) to be specifically enriched in podocytes in human and rodent kidney. Using immunogold labeling in electron microscopy, we were able to demonstrate an enrichment of MAP1B in primary processes. A similar association of MAP1B with the microtubule cytoskeleton was detected in cultured podocytes. Subcellular distribution of MAP1B HC and LC1 was analyzed using a double fluorescent reporter MAP1B fusion protein. Subsequently we analyzed mice constitutively depleted of MAP1B. Interestingly, MAP1B KO was not associated with any functional or structural alterations pointing towards a redundancy of MAP proteins in podocytes. In summary, we established MAP1B as a specific marker protein of the podocyte microtubular cytoskeleton.     

Microtubule-associated proteins 1A and LC2. Two proteins encoded in one messenger RNA.

The deduced amino acid sequence for the filamentous microtubule-associated protein (MAP) 1A, thought to be involved in stabilizing the mature neuronal cytoskeleton, has been determined from a series of overlapping cDNA clones. Though previously described as biochemically and immunologically distinct from MAP1B, we now demonstrate that MAP1A is structurally related to MAP1B, a protein associated with neurite outgrowth and process plasticity. The two MAPs exhibit regional amino acid sequence similarities spanning their potential microtubule binding domains placing both into a new MAP family. The cDNA sequence encoding MAP1A was also found to encode one of its associated light chains (LC) called LC2. Both proteins are found on a single mRNA in the same open reading frame and are translated as a pre-MAP1A/LC2-protein. The topological relationship between MAP1A and LC2 coding sequences is, therefore, identical to that previously shown for MAP1B and LC1 (Hammarback, J. A., Obar, R. A., Hughes, S. M., and Vallee, R. B. (1991) Neuron 7, 129-139). Based on these and earlier results, we conclude that LC1 and LC2 are structurally related polypeptides generated from distinct MAP polyprotein precursors but free to exchange between the two MAPs.     

Interaction of casein kinase 1 delta (CK1delta) with post-Golgi structures, microtubules and the spindle apparatus

Members of the casein kinase 1 family of serine/threonine kinases are highly conserved from yeast to mammals and seem to play an important role in vesicular trafficking, DNA repair, cell cycle progression and cytokinesis. We here report that in interphase cells of various mammalian species casein kinase 1 delta (CK1delta) specifically interacts with the trans Golgi network and cytoplasmic, granular particles that associate with microtubules. Furthermore, at mitosis CK1delta is recruited to the spindle apparatus and the centrosomes in cells, which have been exposed to DNA-damaging agents like etoposide or gammairradiation. In addition, determination of the affinity of CK1delta to different tubulin isoforms in immunoprecipitation-Western analysis revealed a dramatically enhanced complex formation between CK1delta and tubulins from mitotic extracts after introducing DNA damage. The high affinity of CK1delta to the spindle apparatus in DNA-damaged cells and its ability to phosphorylate several microtubule-associated proteins points to a regulatory role of CK1delta at mitosis.     

Identification of novel protein kinase CK1 delta (CK1delta) inhibitors through structure-based virtual screening

In eukaryotes, protein phosphorylation of serine, threonine or tyrosine residues by protein kinases plays an important role in many cellular processes. Members of the protein kinase CK1 family usually phosphorylate residues of serine that are close to other phosphoserine in a consensus motif of pS-X-X-S, and they are implicated in the regulation of a variety of physiological processes as well as in pathologies like cancer and Alzheimer's disease. Using a structure-based virtual screening (SBVS) approach we have identified two anthraquinones as novel CK1delta inhibitors. These amino-anthraquinone analogs (derivatives 1 and 2) are among the most potent and selective CK1delta inhibitors known today (IC(50)=0.3 and 0.6 microM, respectively).     

The Light Chain 1 Subunit of the Microtubule-Associated Protein 1B (MAP1B) Is Responsible for Tiam1 Binding and Rac1 Activation in Neuronal Cells

Microtubule-associated protein 1B (MAP1B) is a neuronal protein involved in the stabilization of microtubules both in the axon and somatodendritic compartments. Acute, genetic inactivation of MAP1B leads to delayed axonal outgrowth, most likely due to changes in the post-translational modification of tubulin subunits, which enhances microtubule polymerization. Furthermore, MAP1B deficiency is accompanied by abnormal actin microfilament polymerization and dramatic changes in the activity of small GTPases controlling the actin cytoskeleton. In this work, we showed that MAP1B interacts with a guanine exchange factor, termed Tiam1, which specifically activates Rac1. These proteins co-segregated in neurons, and interact in both heterologous expression systems and primary neurons. We dissected the molecular domains involved in the MAP1B-Tiam1 interaction, and demonstrated that pleckstrin homology (PH) domains in Tiam1 are responsible for MAP1B binding. Interestingly, only the light chain 1 (LC1) of MAP1B was able to interact with Tiam1. Moreover, it was able to increase the activity of the small GTPase, Rac1. These results suggest that the interaction between Tiam1 and MAP1B, is produced by the binding of LC1 with PH domains in Tiam1. The formation of such a complex impacts on the activation levels of Rac1 confirming a novel function of MAP1B related with the control of small GTPases. These results also support the idea of cross-talk between cytoskeleton compartments inside neuronal cells.     

MAP1A light chain 2 interacts with exchange protein activated by cyclic AMP 1 (EPAC1) to enhance Rap1 GTPase activity and cell adhesion

We have recently demonstrated that light chain 2 (LC2) of the microtubule-associated protein MAP1A interacts with the cyclic AMP (cAMP)-binding domain of exchange protein directly activated by cyclic AMP 1 (EPAC1). In the present study we used a simultaneous expression system and found that LC2 enhances both basal and 8-(4-chloro-phenylthio)-2'-O-methyladenosine-3':5'-cyclic monophosphate (8-CPT-2Me-cAMP)-stimulated Rap1 activation by EPAC1. LC2 is known to stabilize microtubules; therefore we examined whether microtubules enhanced Rap1 activation by LC2. Nocodazole inhibited Rap1 activity in cells transfected with EPAC1 alone but had little effect on Rap1 activity in cells transfected with both EPAC1 and LC2. This indicates that part of the actions of LC2 in enhancing EPAC1 activity may be through stabilization of microtubules. We also found that in cells transfected with LC2, Rap1 was more sensitive to activation by 8-CPT-2Me-cAMP. Moreover, LC2 enhanced the ability of transfected and endogenous EPAC1 to interact with cyclic AMP-agarose, indicating that LC2 elicits conformational changes in the cAMP domain of EPAC1, enhancing its ability to be activated by cyclic AMP. We also found that disruption of the interaction of endogenous EPAC1 and LC2 with antibodies to the cAMP domain of EPAC1 abolished Rap1 activity in PC12 cell lysates, demonstrating the importance of LC2 for EPAC1 activation in these cells. Consistent with a role of EPAC1 in controlling integrin activity, we found that cell adhesion to laminin was enhanced in LC2- and EPAC1-transfected cells stimulated with 8-CPT-2Me-cAMP. LC2 is therefore a biological enhancer of EPAC1 activity toward Rap1 and associated downstream signaling mechanisms.     

Bmcc1s, a novel brain-isoform of Bmcc1, affects cell morphology by regulating MAP6/STOP functions

The BCH (BNIP2 and Cdc42GAP Homology) domain-containing protein Bmcc1/Prune2 is highly enriched in the brain and is involved in the regulation of cytoskeleton dynamics and cell survival. However, the molecular mechanisms accounting for these functions are poorly defined. Here, we have identified Bmcc1s, a novel isoform of Bmcc1 predominantly expressed in the mouse brain. In primary cultures of astrocytes and neurons, Bmcc1s localized on intermediate filaments and microtubules and interacted directly with MAP6/STOP, a microtubule-binding protein responsible for microtubule cold stability. Bmcc1s overexpression inhibited MAP6-induced microtubule cold stability by displacing MAP6 away from microtubules. It also resulted in the formation of membrane protrusions for which MAP6 was a necessary cofactor of Bmcc1s. This study identifies Bmcc1s as a new MAP6 interacting protein able to modulate MAP6-induced microtubule cold stability. Moreover, it illustrates a novel mechanism by which Bmcc1 regulates cell morphology.     

Catalytic activity of protein kinase CK1 delta (casein kinase 1delta) is essential for its normal subcellular localization

Mammalian casein kinase 1delta (CK1delta) is a homologue of the S. cerevisiae Hrr25p protein kinase. Hrr25p is involved in regulating diverse events including vesicular trafficking, gene expression, DNA repair, and chromosome segregation. In contrast to Hrr25p, little is known about the function, regulation, or subcellular localization of CK1delta. In the present study, we show that CK1delta in mammalian cells is mainly cytoplasmic and enriched within the Golgi and/or ER-Golgi transport vesicles, consistent with a role in vesicular trafficking. Transient expression of green fluorescent protein (GFP)- or FLAG peptide-tagged CK1delta showed localization similar to that of the endogenous CK1delta. GFP-CK1delta was also enriched at the centrosomes in interphase cells. Strikingly, two inactive mutant CK1delta proteins (K38M and T176I) showed almost exclusive nuclear staining, suggesting that protein kinase activity is required for normal localization of CK1delta and prevention of nuclear accumulation. The nuclear export inhibitor leptomycin B promoted nuclear enrichment of CK1delta indicating that nuclear localization of CK1delta occurs physiologically. Both endogenous CK1delta and GFP-CK1delta are enriched on the spindle poles in mitotic cells, consistent with a role in regulating spindle formation. Localization is a property of the protein kinase domain and is independent of the C-terminal noncatalytic domain. These data are consistent with roles for CK1delta in mammalian cells analogous to those of its yeast counterparts.     

The MAP1 family of microtubule-associated proteins

MAP1-family proteins are classical microtubule-associated proteins (MAPs) that bind along the microtubule lattice. The founding members, MAP1A and MAP1B, are predominantly expressed in neurons, where they are thought to be important in the formation and development of axons and dendrites. Mammalian genomes usually contain three family members, MAP1A, MAP1B and a shorter, more recently identified gene called MAP1S. By contrast, only one family member, Futsch, is found in Drosophila. After their initial expression, the MAP1A and MAP1B polypeptides are cleaved into light and heavy chains, which are then assembled into mature complexes together with the separately encoded light chain 3 subunit (LC3). Both MAP1A and MAP1B are well known for their microtubule-stabilizing activity, but MAP1 proteins can also interact with other cellular components, including filamentous actin and signaling proteins. Furthermore, the activity of MAP1A and MAP1B is controlled by upstream signaling mechanisms, including the MAP kinase and glycogen synthase kinase-3 β pathways.     

Lack of communication between LC2 light chain and the SH1 region of myosin S-1 studied by 19F-NMR

Myosin subfragment-1 (S-1) which contains the LC2 light chain has been labelled with fluorine to allow an 19F-NMR study of the coupling and energetics of structural changes in the myosin head. Two fluorine-containing reagents, N-4-(trifluoromethyl)phenyl iodoacetamide and N-3,5-di(trifluoromethyl)phenyl iodoacetamide, have been used to label the myosin heavy chain at the unusually reactive sulfhydryl-1 (SH1) position. The chemical shift of both reagents on S-1 is sensitive to a structural transition in the region of SH1 which occurs upon increasing the temperature from 0 degrees C to 35 degrees C. The midpoint of the transition in both papain and chymotryptic S-1 is at approximately 11 degrees C at pH 7 (0.1 M CKl). The temperature dependence of the chemical shift may be fit assuming a two-state equilibrium where delta G degree' (T) = 101-110T +0.386 T2 (where T is the temperature in Kelvin). Both delta H degree' (T) and delta S degree' (T) have a small temperature dependence from 0 to 35 degrees C: at 20 degrees C, delta H degree' (T) = -33 kcal/mol. delta S degree' (T) = -116 e.u. and delta Cp = -226 cal/mol per deg (pH 7.0, 0.1 M KCl). The NMR data indicate that the presence of the LC2 light chain in papain S-1 does not modify the structure of S-1 in the vicinity of SH1, nor does it modify the energetics of the structural transition from that seen in its absence with chymotryptic S-1. The presence of calcium which is bound by the LC2 of papain S-1 also does not alter the energetics of the transition. Thus it would appear that the LC2 light chain (on myosin S-1) does not participate in the two-state transition, nor does it interact strongly with regions of the heavy chain which participate in the transition.     

The MAP1B-LC1/UBE2L3 complex catalyzes degradation of cell surface CaV2.2 channels

We reported recently a new mechanism by which the neuronal N-type Ca²⁺ (Ca_V2.2) channel expression may be regulated by ubiquitination. This mechanism involves the interaction between the channel and the light chain (LC1) of the microtubule associated protein B (MAP1B). We also showed that MAP1B-LC1 could interact with the ubiquitin-conjugating E2 enzyme UBE2L3 and that the ubiquitination/degradation mechanism triggered by MAP1B-LC1 could be prevented by inhibiting the ubiquitin-proteasome proteolytic pathway. We now report that MAP1B-LC1 can interact with the 2 main variants of the Ca_V2.2 channels (Ca_V2.2e37a and Ca_V2.2e37b) and that the MAP1B-LC1-mediated regulation most likely involves an internalization of the channels via a dynamin and clathrin-dependent pathway. In addition, here we propose that this novel mechanism of Ca_V channel regulation might be conserved among N-type and P/Q-type channels.     

The MAP1B-LC1/UBE2L3 complex catalyzes degradation of cell surface CaV2.2 channels

We reported recently a new mechanism by which the neuronal N-type Ca²⁺ (Ca_V2.2) channel expression may be regulated by ubiquitination. This mechanism involves the interaction between the channel and the light chain (LC1) of the microtubule associated protein B (MAP1B). We also showed that MAP1B-LC1 could interact with the ubiquitin-conjugating E2 enzyme UBE2L3 and that the ubiquitination/degradation mechanism triggered by MAP1B-LC1 could be prevented by inhibiting the ubiquitin-proteasome proteolytic pathway. We now report that MAP1B-LC1 can interact with the 2 main variants of the Ca_V2.2 channels (Ca_V2.2e37a and Ca_V2.2e37b) and that the MAP1B-LC1-mediated regulation most likely involves an internalization of the channels via a dynamin and clathrin-dependent pathway. In addition, here we propose that this novel mechanism of Ca_V channel regulation might be conserved among N-type and P/Q-type channels.     

A role for Tctex-1 (DYNLT1) in controlling primary cilium length

The microtubule motor complex cytoplasmic dynein is known to be involved in multiple processes including endomembrane organization and trafficking, mitosis, and microtubule organization. The majority of studies of cytoplasmic dynein have focused on the form of the motor that is built around the dynein-1 heavy chain. A second isoform, dynein heavy chain-2, and its specifically associated light intermediate chain, LIC3 (D2LIC), are known to be involved in the formation and function of primary cilia. We have used RNAi in human epithelial cells to define the cytoplasmic dynein subunits that function with dynein heavy chain 2 in primary cilia. We identify the dynein light chain Tctex-1 as a key modulator of cilia length control; depletion of Tctex-1 results in longer cilia as defined by both acetylated tubulin labeling of the axoneme and Rab8a labeling of the cilia membrane. Suppression of dynein heavy chain-2 causes concomitant loss of Tctex-1 and this correlates with an increase in cilia length. Compared to individual depletions, double siRNA depletion of DHC2 and Tctex-1 causes an even greater increase in cilia length. Our data show that Tctex-1 is a key regulator of cilia length and most likely functions as part of dynein-2.     

Heterotypic complex formation between subunits of microtubule-associated proteins 1A and 1B is due to interaction of conserved domains

The microtubule-associated proteins MAP1A and MAP1B are related but distinct multi-subunit protein complexes that consist of heavy and light chains. The predominant forms of these complexes are homotypic, i.e. they consist of a MAP1A heavy chain associated with MAP1A light chains or a MAP1B heavy chain associated with MAP1B light chains, respectively. In addition, MAP1A and MAP1B can exchange subunits and form heterotypic complexes consisting of a MAP1A heavy chain associated with MAP1B light chains which might play a role in a transition period of neuronal differentiation. Here we extend previous findings by confirming that heterotypic MAP1B heavy chain-MAP1A light chain complexes also exist in the developing murine brain. We show that these complexes form through interaction of homologous domains conserved in heavy and light chains of MAP1A and MAP1B. Likewise, conserved domains of the MAP1A and MAP1B light chains account for formation of light chain heterodimers. By yeast 2-hybrid analysis we located the light chain binding domain on the heavy chain to amino acids 211-508, thereby defining a new functional subdomain.